Methods and compositions for sample processing

Abstract

Provided herein are methods and compositions for processing a target nucleic acid sequence. The methods and compositions provided herein comprise a detergent, a solubilizer, and a cyclodextrin configured to stabilize an enzyme and nucleic acids. The processed target nucleic acid sequence can be further contacted with a reaction mixture used in a nucleic acid amplification (e.g., isothermal amplification). A processed target nucleic acid sample can be contacted with a stabilization buffer configured to stabilize one or more enzymes in a nucleic acid amplification. The processed target nucleic acid sample can be further contacted with an amplification buffer configured to increase a rate of amplification during a nucleic acid amplification (e.g., isothermal amplification).

Description

METHODS AND COMPOSITIONS FOR SAMPLE PROCESSINGCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 580,018, filed on September 1, 2023, the entire content of which is entirely incorporated herein by reference.BACKGROUND OF THE INVENTION

[0002] Nucleic acid amplification techniques such as polymerase chain reaction (PCR) and various isothermal amplification techniques have become an integral part of nucleic acid-based diagnostics and research techniques. Samples containing target nucleic acid molecules need to be processed before being amplified in a nucleic acid amplification assay. For example, the target nucleic acid molecules need to be extracted. Optimization of sample processing and preparation can improve the efficiency, precision, and yield in nucleic acid amplification techniques.SUMMARY OF THE INVENTION

[0003] Recognized herein is a need for improved methods and compositions for processing and preparing a sample for a nucleic acid amplification technique. The methods and compositions described herein can be used for processing a sample for downstream applications such as isothermal amplifications with improvements in precision, efficiency, and / or yield of target nucleic acid molecule products. The sample processing, stabilization, and amplification methods and compositions provided herein can shorten the time to result value to be less than 30 minutes.

[0004] In some aspects, the present disclosure provides a composition for sample processing comprising: a detergent, a solubilizer, and a cyclodextrin, wherein the composition is configured to stabilize an enzyme during a nucleic acid amplification, and wherein the composition is configured to reduce and / or eliminate activity of a degrading nuclease.

[0005] In some embodiments, the composition is configured to stabilize nucleic acids during the nucleic acid amplification. In some embodiments, the enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof. In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.WSGR Docket No. 52459-726.601

[0006] In some embodiments, the solubilizer is a non-ionic surfactant. In some embodiments, the solubilizer is a polysorbate, octylphenoxypolyethoxyethanol, 2-[4-(2,4,4-trimethylpentan-2- yl)phenoxy]ethanol, or a secondary alcohol ethoxylate. In some embodiments, the solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof. In some embodiments, the detergent is part of a lysis buffer. In some embodiments, the solubilizer and the cyclodextrin are part of a recovery buffer. In some embodiments, the lysis buffer and the recovery buffer are in a same mixture.

[0007] In some aspect, the present disclosure provides a composition for sample processing comprising a buffer comprising: a detergent, a solubilizer, and a cyclodextrin, wherein the composition is configured to stabilize an enzyme during a nucleic acid amplification, and wherein the composition is configured to inactivate a degrading nuclease.

[0008] In some embodiments, the composition is configured to stabilize nucleic acids during the nucleic acid amplification. In some embodiments, the enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof. In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof. In some embodiments, the solubilizer and the cyclodextrin are configured to shorten a cycle threshold value or a time to result value in the nucleic acid amplification compared to a cycle threshold value or a time to result value in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually.

[0009] In some embodiments, the cycle threshold value is at most 40 or the time to result value is at most 15 minutes. In some embodiments, the solubilizer and the cyclodextrin are configured to decrease a coefficient of variation. In some embodiments, the solubilizer and the cyclodextrin are configured to lower a limit of detection. In some embodiments, the degrading nuclease is a ribonuclease. In some embodiments, the lysis buffer has a pH value of 2 to 9. In some embodiments, the lysis buffer further comprises a chelating agent. In some embodiments, the chelating agent is deferiprone, ethylenediamine, 1,10-Phenanthroline, oxalic acid, pentetic acid, deferasirox, deferoxamine, deferoxamine mesylate, or N,N,N',N'-tetrakis(2-pyridinylmethyl)- 1,2-ethanediamine (TPEN).

[0010] In some embodiments, the lysis buffer further comprises a reducing agent. In some embodiments, the reducing agent is oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-WSGR Docket No. 52459-726.601 tetraacetic acid (BAPTA), or tetrahydropyran (THP). In some embodiments, the lysis buffer comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2- carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP). In some embodiments, a final concentration of EGTA in the lysis buffer in the presence of a sample is about 0.1 millimolar (mM) to 10 mM, a final concentration of EDTA in the lysis buffer in the presence of a sample is about 0.1 mM to 5 mM, a final concentration of TCEP in the lysis buffer in the presence of a sample is about 1 mM to 20 mM, or a final concentration of Tris in the lysis buffer in the presence of a sample is about 1 mM to 60 mM.

[0011] In some embodiments, the composition further comprises an agent capable of reducing a disulfide bond. In some embodiments, the agent capable of reducing the disulfide bond comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2-mercaptoehtanol (0ME).

[0012] In some embodiments, the detergent is present in the composition mixed with a sample at a final concentration that is effective for lysing cells. In some embodiments, the cyclodextrin is present in the composition mixed with a sample at a final concentration that is effective for isolating the detergent within the composition. In some embodiments, the detergent is configured to form a complex with the solubilizer and / or the cyclodextrin to stabilize the enzyme. In some embodiments, the cyclodextrin is configured to increase the efficiency of forming the complex. In some embodiments, the cyclodextrin has a higher binding affinity toward the detergent than a binding affinity of the solubilizer. In some embodiments, the final concentration of the detergent is about 0.1% to 10% w / v (g of solute / 100 mL of solution). In some embodiments, the final concentration of the cyclodextrin is about 0.1 mM to 70 mM. In some embodiments, the cyclodextrin comprises hydroxypropyl 0-cyclodextrin, hydroxypropyl y- cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a- cyclodextrin hydrate, monopropanediamino-0-cyclodextrin, 6-O-alpha-D-Maltosyl-0- cyclodextrin, 2,6-Di-O-methyl-0-cyclodextrin, hydroxyethyl-0-cyclodextrin, 3A-amino-3A- deoxy-(2AS,3AS)-0-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, an anionic cyclodextrin, or any combination thereof.WSGR Docket No. 52459-726.601

[0013] In some embodiments, the solubilizer is present in the composition mixed with a sample at a final concentration of about 0.1% to 50% w / v. In some embodiments, the final concentration of the solubilizer is effective for forming micelles comprising the detergent. In some embodiments, the recovery buffer comprises a salt. In some embodiments, the recovery buffer does not comprise a salt. In some embodiments, the recovery buffer comprises a pH buffer. In some embodiments, the recovery buffer does not comprise a pH buffer. In some embodiments, the lysis buffer is lyophilized. In some embodiments, the recovery buffer is lyophilized.

[0014] In some embodiments, the composition further comprises a sample. In some embodiments, the sample is a biological sample. In some embodiments, the biological sample comprises a target nucleic acid molecule subject to sample processing.

[0015] In some embodiments, the composition further comprises a reaction mixture for nucleic acid amplification. In some embodiments, the reaction mixture is lyophilized. In some embodiments, the reaction mixture comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, or a probe. In some embodiments, the composition is configured to stabilize enzymatic activity of the thermostable enzyme for use during a nucleic acid amplification. In some embodiments, the thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a B st 2.0 polymerase, a B st 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and any mutants thereof. In some embodiments, the dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP. In some embodiments, a concentration of the dNTPs in the reaction mixture is about 40 micromolar (pM) to 5000 pM. In some embodiments, the primer is at least 4 nucleotides in length.

[0016] In some aspects, the present disclosure provides a method of processing a sample, the method comprising mixing a sample with the lysis buffer described herein.

[0017] In some embodiments, the method further comprises mixing the sample with the recovery buffer described herein.

[0018] In some aspects, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a lysis buffer comprising a detergent; and (b) contacting the sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing the sample to generate a processed sample in a mixture comprising the detergent, the solubilizer, and the cyclodextrin.WSGR Docket No. 52459-726.601

[0019] In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

[0020] In some embodiments, the sample is a biological sample. In some embodiments, the sample is a purified sample. In some embodiments, (a) and (b) occur simultaneously. In some embodiments, contacting the sample in (a) and (b) is performed concurrently in the same mixture. In some embodiments, the method further comprises incubating the sample at room temperature for a duration of time. In some embodiments, the method further comprises heating the sample at a constant temperature for a period of time. In some embodiments, the method further comprises heating the sample at a cyclic temperature for a period of time. In some embodiments, the method further comprises sonicating the sample. In some embodiments, sonicating the sample occurs prior to, subsequent to, or concurrent to heating the sample.

[0021] In some aspects, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a lysis buffer comprising a detergent; (b) incubating the sample at a first temperature or temperature range for a first time period; (c) heating the sample at a second temperature or temperature range for a second time period; and (d) contacting the sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing the sample to generate a processed sample in a mixture comprising the detergent, the solubilizer and the cyclodextrin.

[0022] In some embodiments, the heating the sample in (c) further comprises heating the sample to the second temperature, cooling down the sample, and heating the sample to the second temperature after cooling down. In some embodiments, the method further comprises sonicating the sample. In some embodiments, the sonicating the sample is performed prior to, subsequent to, or concurrent to heating the sample. In some embodiments, the method further comprises bead beating the sample. In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

[0023] In some embodiments, the sample is a biological sample. In some embodiments, the sample is a purified sample. In some embodiments, the first temperature or temperature range is about 2°C to 25°C. In some embodiments, the second temperature is about 60°C to 100°C. InWSGR Docket No. 52459-726.601 some embodiments, the first time period is at least about 1 minute to at least about 48 hours. In some embodiments, the second time period is at least about 1 to at least about 10 minutes.

[0024] In some embodiments, the lysis buffer further comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)- 1,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

[0025] In some embodiments, the method further comprises, subsequent to contacting the sample with the recovery buffer, contacting the processed sample with a reaction mixture. In some embodiments, the reaction mixture comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, or a probe. In some embodiments, the reaction mixture stabilizes enzymatic activity of the thermostable enzyme for use during a nucleic acid amplification. In some embodiments, the thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a B st 2.0 polymerase, a B st 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and any mutants thereof. In some embodiments, the dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP. In some embodiments, a concentration of the dNTPs in the reaction mixture is about 40 pM to 5000 pM. In some embodiments, the primer is at least 4 nucleotides in length.

[0026] In some embodiments, the method further comprises subjecting the processed sample mixed with the reaction mixture to a nucleic acid amplification. In some embodiments, the nucleic acid amplification comprises polymerase chain reaction (PCR) or isothermal amplification. In some embodiments, the nucleic acid amplification comprises thermocycling the processed sample. In some embodiments, the nucleic acid amplification comprises keeping the processed sample at a constant temperature for amplification. In some embodiments, the method further comprising, prior to (a), obtaining the sample from a subject. In some embodiments, the subject has or is suspected of having a disease, a condition, or an infection. In some embodiments, the sample comprises one or more different target nucleic acid molecules.

[0027] In some embodiments, the sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, a lymph sample, raw milk, pasteurized and / orWSGR Docket No. 52459-726.601 homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof.

[0028] In some embodiments, a time from obtaining the sample to generating the processed sample is equal to or less than about 30 min, 25 min, 20 min, 15 min, 10 min, 5 min, 4 min, 3 min, 2 min, 1 min or less. In some embodiments, a concentration of the one or more different target nucleic acid molecules is at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% or more compared to a concentration of one or more different target nucleic acid molecules of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually.

[0029] In some aspects, the present disclosure provides a kit for sample processing, the kit comprising a lysis buffer comprising a detergent, a recovery buffer comprising a solubilizer and a cyclodextrin, and an instruction for use.

[0030] In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof. In some embodiments, the kit further comprises a reagent for nucleic acid amplification comprising a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), or a primer. In some embodiments, the thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and any mutants thereof. In some embodiments, the dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP. In some embodiments, a concentration of the dNTPs in a reaction mixture is about 40 pM to 5000 pM. In some embodiments, the primer is at least 4 nucleotides in length. In some embodiments, the kit further comprises a probe for detecting an amplification product generated using the kit. In some embodiments, the lysis buffer, the recovery buffer or the reagent is lyophilized. In some embodiments, the recovery buffer further comprises a cucurbituril. In some embodiments, the cucurbituril is cucurbit[n]uril, wherein n is an integer of 5, 6, 7, 8, or 10.

[0031] In some embodiments, the reaction mixture comprises an excipient. In some embodiments, the excipient comprises one or more reagents selected from the group consisting of a Tris, potassium phosphate, sodium chloride, ethylenediaminetetraacetic acid (EDTA),WSGR Docket No. 52459-726.601 potassium chloride, nonoxynol-9, trehalose, dextran, polysucrose 400, and a cyclodextrin. In some embodiments, the cyclodextrin comprises hydroxypropyl P-cyclodextrin, hydroxypropyl y- cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a- cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P- cyclodextrin, 2,6-Di-O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A- deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, an anionic cyclodextrin, or any combination thereof. In some embodiments, a final concentration of Tris in the excipient in the presence of the sample is about 0.001 molar (M) to 1.0 M; a final concentration of sodium chloride and / or potassium chloride in the presence of the sample is about 0.0001 M to 0.25 M; a final concentration of EDTA in the excipient in the presence of the sample is about 0.00001 M to 0.1 M; a final concentration of nonoxynol-9 in the excipient in the presence of the sample is about 0.01% v / v to 2.0% v / v; a final concentration of trehalose in the excipient in the presence of the sample is about .001 M to 2.0 M; a final concentration of dextran in the excipient in the presence of the sample is about 0.1% w / v to 10% w / v; a final concentration of poly sucrose 400 in the excipient in the presence of the sample is about 0.01% w / v to 5.0% w / v; and / or a final concentration of the cyclodextrin in the excipient in the presence of the sample is about 0.001 M to 5.0 M.

[0032] In some embodiments, the excipient further comprises an additional reagent. In some embodiments, the additional reagent comprises a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof. In some embodiments, the additional reagent is configured to stabilize an enzyme. In some embodiments, the additional reagent is configured to lower a Cq value of a nucleic acid amplification.

[0033] In some embodiments, the composition further comprises a sample stabilization buffer. In some embodiments, the sample stabilization buffer comprises one or more reagents selected from the group consisting of a collapse modifier, a protein stabilizer, and a glass transition modifier. In some embodiments, the sample stabilization buffer comprises at least one salt. In some embodiments, the sample stabilization buffer comprises a cyclodextrin. In some embodiments, the sample stabilization buffer is configured to reconstitute a lyophilized sample. In some embodiments, the stabilization buffer comprises at least one reducing agent. In some embodiments, the at least one reducing agent is oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, l,2-bis(o-WSGR Docket No. 52459-726.601 aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP), or any combination thereof.

[0034] In some embodiments, a total time to perform (a) and (b) is at most 1 min, at most 50 seconds, at most 40 seconds, or at most 20 seconds. In some embodiments, a time for processing the sample is a time period from the contacting of (a) to contacting the processed sample with a reaction mixture, wherein the time period is at most 20 seconds. In some embodiments, the nucleic acid amplification generates an amplified processed sample. In some embodiments, a time period of the nucleic acid amplification to generate the amplified processed sample is at most 5 minutes. In some embodiments, the method of processing the sample does not comprise heating the sample.

[0035] In some aspects, the present disclosure provides a composition for sample amplification comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer, wherein the composition is configured to increase a rate of amplification during a nucleic acid amplification.

[0036] In some embodiments, the composition is configured to stabilize an enzyme during a nucleic acid amplification. In some embodiments, the enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof. In some embodiments, the reverse transcriptase is an avian myeloblastosis virus (AMV) reverse transcriptase or a murine leukemia virus (MMLV) reverse transcriptase. In some embodiments, the nonionic surfactant is nonoxynol-9. In some embodiments, a final concentration of the cyclodextrin in the composition in the presence of a sample is 0.01% v / v to 2.0% v / v. In some embodiments, the cyclodextrin comprises the cyclodextrin comprises hydroxypropyl P-cyclodextrin, hydroxypropyl y- cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a- cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P- cyclodextrin, 2,6-Di-O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A- deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, or any combination thereof. In some embodiments, a final concentration of the cyclodextrin in the composition in the presence of a sample is about 0.001 M to 10 M.

[0037] In some embodiments, the sucrose / epichlorohydrin polymer is polysucrose 400. In some embodiments, a final concentration of the sucrose / epichlorohydrin polymer in the composition in the presence of a sample is about 0.001% to 5% w / v (g of solute / 100 mL of solution). In some embodiments, the composition further comprises at least one salt. In some embodiments, a final concentration of the at least one salt in the composition in the presence of a sample is aboutWSGR Docket No. 52459-726.6010.001 molar (M) to 10 M. In some embodiments, the at least one salt is sodium chloride, potassium phosphate, potassium chloride, or any combination thereof. In some embodiments, the composition comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10- Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, 1,2- bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP). In some embodiments, a final concentration of EDTA in the composition in the presence of a sample is about 0.01 millimolar (mM) to 10 mM, and / or a final concentration of Tris in the composition in the presence of a sample is about 0.1 mM to 25 mM.

[0038] In some embodiments, the composition further comprises an agent capable of reducing a disulfide bond. In some embodiments, the agent capable of reducing the disulfide bond comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2-mercaptoehtanol (PME).

[0039] In some embodiments, the composition further comprises at least one sugar and / or sugar alcohol. In some embodiments, the at least one sugar and / or sugar alcohol comprises sucrose, lactose, trehalose, dextran, erythritol, arabitol, xylitol, sorbitol, mannitol, or any combination thereof. In some embodiments, a final concentration of the at least one sugar and / or sugar alcohol is about 0.001 M to 10 M or about 0.1% to 10% w / v (g of solute / 100 mL of solution).

[0040] In some embodiments, the composition further comprises an additional reagent. In some embodiments, the additional reagent comprises a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof. In some embodiments, the composition is lyophilized.

[0041] In some embodiments, the composition further comprises a sample. In some embodiments, the sample is a biological sample. In some embodiments, the biological sample comprises a target nucleic acid molecule subject to sample processing.

[0042] In some embodiments, the composition further comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, a probe, or any combination thereof. In some embodiments, the composition is configured to stabilize enzymatic activity of the thermostable enzyme for use during the nucleic acid amplification. In some embodiments, the thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0WSGR Docket No. 52459-726.601 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exopolymerase, an OmniTaq 2 LA DNA polymerase, a IsoFast™ Bst, and any mutants thereof. In some embodiments, the dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP. In some embodiments, a concentration of the dNTPs in the composition is about 40 micromolar (pM) to 5000 pM. In some embodiments, the primer is at least 4 nucleotides in length. In some embodiments, the probe is at least 15 nucleotides in length.

[0043] In some aspects, the present disclosure provides a composition comprising: a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer; and a sample stabilization buffer configured to stabilize an enzyme in a nucleic acid amplification.

[0044] In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the solubilizer is a non-ionic surfactant. In some embodiments, the solubilizer is a polysorbate, octylphenoxypolyethoxyethanol, 2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol, or a secondary alcohol ethoxylate. In some embodiments, the solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof. In some embodiments, the detergent is part of a lysis buffer. In some embodiments, the solubilizer and the cyclodextrin are part of a recovery buffer. In some embodiments, the lysis buffer and the recovery buffer are in the sample processing buffer as the same mixture. In some embodiments, the solubilizer and the cyclodextrin are configured to shorten a cycle threshold value or a time to result value in the nucleic acid amplification compared to a cycle threshold value or a time to result value in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually. In some embodiments, the cycle threshold value is at most 40 or the time to result value is at most 15 minutes.

[0045] In some embodiments, the solubilizer and the cyclodextrin are configured to decrease a coefficient of variation. In some embodiments, the solubilizer and the cyclodextrin are configured to lower a limit of detection.

[0046] In some embodiments, the lysis buffer further comprises a chelating agent. In some embodiments, the chelating agent is deferiprone, ethylenediamine, 1,10-Phenanthroline, oxalic acid, pentetic acid, deferasirox, deferoxamine, deferoxamine mesylate, or N,N,N',N'-tetrakis(2- pyridinylmethyl)-l,2-ethanediamine (TPEN). In some embodiments, the lysis buffer further comprises a reducing agent. In some embodiments, the reducing agent is oxalic acid, formicWSGR Docket No. 52459-726.601 acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, 1,2- bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP). In some embodiments, the lysis buffer comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)- 1,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

[0047] In some embodiments, the composition described herein comprises a final concentration of EGTA in the lysis buffer in the presence of a sample is about 0.1 millimolar (mM) to 10 mM, a final concentration of EDTA in the lysis buffer in the presence of a sample is about 0.1 mM to 5 mM, a final concentration of TCEP in the lysis buffer in the presence of a sample is about 1 mM to 20 mM, or a final concentration of Tris in the lysis buffer in the presence of a sample is about 1 mM to 60 mM.

[0048] In some embodiments, the sample processing buffer further comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2-mercaptoehtanol (0ME). In some embodiments, the detergent is present in the sample processing buffer mixed with a sample at a final concentration that is effective for lysing cells. In some embodiments, the cyclodextrin is present in the sample processing buffer mixed with a sample at a final concentration that is effective for isolating the detergent within the composition. In some embodiments, the detergent is configured to form a complex with the solubilizer and / or the cyclodextrin to stabilize the enzyme. In some embodiments, the cyclodextrin is configured to increase the efficiency of forming the complex. In some embodiments, the final concentration of the detergent is about 0.1% to 10% w / v (g of solute / 100 mL of solution). In some embodiments, the final concentration of the cyclodextrin is about 0.1 mM to 70 mM.

[0049] In some embodiments, the cyclodextrin comprises hydroxypropyl P-cyclodextrin, hydroxypropyl y-cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy- (2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D- Maltosyl-P-cyclodextrin, 2,6-Di-O-methyl-p-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A- amino-3A-deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y- cyclodextrin hydrate, or any combination thereof.WSGR Docket No. 52459-726.601

[0050] In some embodiments, the solubilizer is present in the composition mixed with a sample at a final concentration of about 0.1% to 50% w / v. In some embodiments, the final concentration of the solubilizer is effective for forming micelles comprising the detergent. In some embodiments, the recovery buffer comprises a salt. In some embodiments, the recovery buffer comprises a pH buffer. In some embodiments, the recovery buffer does not comprise a pH buffer. In some embodiments, the sample processing buffer is lyophilized. In some embodiments, the nonionic surfactant of the sample amplification buffer is nonoxynol-9. In some embodiments, a final concentration of the cyclodextrin in the sample amplification buffer in the presence of a sample is 0.01% v / v to 2.0% v / v. In some embodiments, the cyclodextrin comprises the cyclodextrin comprises hydroxypropyl p-cyclodextrin, hydroxypropyl y- cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a- cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P- cyclodextrin, 2,6-Di-O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A- deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, or any combination thereof. In some embodiments, a final concentration of the cyclodextrin in the sample amplification buffer in the presence of a sample is about 0.001 M to 10 M.

[0051] In some embodiments, the sucrose / epichlorohydrin polymer is polysucrose 400. In some embodiments, a final concentration of the sucrose / epichlorohydrin polymer in the composition in the presence of a sample is about 0.001% to 5% w / v (g of solute / 100 mL of solution). In some embodiments, the sample amplification buffer further comprises at least one salt. In some embodiments, a final concentration of the at least one salt in the composition in the presence of a sample is about 0.001 molar (M) to 10 M. In some embodiments, the at least one salt is sodium chloride, potassium phosphate, potassium chloride, or any combination thereof.

[0052] In some embodiments, the sample amplification buffer further comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)- 1,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP). In some embodiments, a final concentration of EDTA in the composition in the presence of a sample is about 0.01 millimolar (mM) to 10 mM, and / or a final concentration of Tris in the composition in the presence of a sample is about 0.1 mM to 60WSGR Docket No. 52459-726.601 mM. In some embodiments, the sample amplification buffer further comprises an agent capable of reducing a disulfide bond. In some embodiments, the agent capable of reducing the disulfide bond comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2- mercaptoehtanol (0ME). In some embodiments, the sample amplification buffer further comprises at least one sugar and / or sugar alcohol. In some embodiments, the at least one sugar and / or sugar alcohol comprises sucrose, lactose, trehalose, dextran, erythritol, arabitol, xylitol, sorbitol, mannitol, or any combination thereof. In some embodiments, a final concentration of the at least one sugar and / or sugar alcohol is about 0.001 M to 10 M or about 0.1% to 10% w / v (g of solute / 100 mL of solution).

[0053] In some embodiments, the sample amplification buffer further comprises an additional reagent. In some embodiments, the additional reagent comprises a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof. In some embodiments, the sample amplification buffer is lyophilized.

[0054] In some embodiments, the sample amplification buffer further comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, a probe, or any combination thereof. In some embodiments, the sample amplification buffer is configured to stabilize enzymatic activity of the thermostable enzyme for use during the nucleic acid amplification. In some embodiments, the thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, a IsoFast™ Bst, and any mutants thereof. In some embodiments, the dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP. In some embodiments, a concentration of the dNTPs in the composition is about 40 micromolar (pM) to 5000 pM. In some embodiments, the primer is at least 4 nucleotides in length. In some embodiments, the probe is at least 15 nucleotides in length. In some embodiments, the sample stabilization buffer comprises one or more reagents selected from the group consisting of a collapse modifier, a protein stabilizer, and a glass transition modifier.

[0055] In some embodiments, the sample stabilization buffer comprises at least one salt. In some embodiments, the sample stabilization buffer comprises a cyclodextrin. In some embodiments, the sample stabilization buffer is configured to reconstitute a lyophilized sample. In some embodiments, the stabilization buffer comprises at least one reducing agent. In some embodiments, the at least one reducing agent is oxalic acid, formic acid, lithium aluminumWSGR Docket No. 52459-726.601 hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, l,2-bis(o- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP), or any combination thereof.

[0056] In some embodiments, the composition further comprises a sample. In some embodiments, the sample is a biological sample. In some embodiments, the biological sample comprises a target nucleic acid molecule subject to sample processing. In some embodiments, the enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof.

[0057] In some aspects, the present disclosure provides a method of amplifying a sample, the method comprising: (a) contacting the sample with a sample processing buffer to generate a processed sample; (b) contacting the processed sample with a sample amplification buffer to provide a condition for a nucleic acid amplification; and (c) subjecting the processed sample to the nucleic acid amplification, and wherein prior to the contacting of (b), the sample processing buffer is not removed.

[0058] In some embodiments, the method further comprises contacting the sample with a sample stabilization buffer for stabilizing an enzyme in the nucleic acid amplification. In some embodiments, the method further comprises, prior to (c), contacting the sample with the sample stabilization buffer. In some embodiments, the sample stabilization buffer is in a same mixture as the sample amplification buffer. In some embodiments, the sample stabilization buffer is contacted with the sample after contacting the sample with the sample amplification buffer. In some embodiments, the method does not comprise heating the sample. In some embodiments, the sample processing buffer comprises a lysis buffer and / or a recovery buffer. In some embodiments, the lysis buffer comprises a detergent. In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the recovery buffer comprises a solubilizer and a cyclodextrin. In some embodiments, a processing time for the sample is a time period from the contacting of (a) and the contacting of (b), wherein the time period is at most 20 seconds. In some embodiments, the nucleic acid amplification comprises polymerase chain reaction (PCR) or isothermal amplification. In some embodiments, the nucleic acid amplification comprises thermocycling the sample. In some embodiments, the nucleic acid amplification generates an amplified sample. In some embodiments, a time period from the contacting of (a) to the amplified sample is at most 5 minutes.WSGR Docket No. 52459-726.601

[0059] In some embodiments, the sample is a biological sample. In some embodiments, the biological sample comprises one or more different target nucleic acid molecules. In some embodiments, a concentration of the one or more different target nucleic acid molecules is at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% greater than a concentration of one or more different target nucleic acid molecules of an otherwise identical sample processed by a sample processing buffer alone. In some embodiments, the sample processing buffer further comprises a cucurbituril.

[0060] In some aspects, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a sample processing buffer to generate a processed sample; (b) contacting the processed sample with a sample amplification buffer to provide a condition for a nucleic acid amplification; and (c) subjecting the processed sample to the nucleic acid amplification in the sample amplification buffer, wherein a time period from contacting in (a) to generating the processed sample prior to contacting with the sample amplification buffer is (i) no more than a time for pipetting the sample processing buffer into the sample to mix the sample processing buffer and the sample or is (ii) at most 1 min, at most 50 seconds, at most 40 seconds, or at most 20 seconds.

[0061] In some embodiments, the sample processing buffer comprises a lysis buffer and / or a recovery buffer. In some embodiments, the lysis buffer comprises a detergent. In some embodiments, the detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the recovery buffer comprises a solubilizer and a cyclodextrin. In some embodiments, the sample processing buffer comprises a detergent, a solubilizer, and a cyclodextrin, wherein the sample processing buffer is configured to stabilize an enzyme during a nucleic acid amplification, and wherein the sample processing buffer is configured to reduce and / or eliminate activity of a degrading nuclease. In some embodiments, the sample processing buffer is the composition described herein. In some embodiments, the sample amplification buffer comprises an excipient. In some embodiments, prior to (b), the sample processing buffer is not removed. In some embodiments, the sample amplification buffer comprises the reaction mixture described herein, or the sample amplification buffer is the composition described herein. In some embodiments, the method does not comprise heating the sample.

[0062] In some embodiments, the sample is a biological sample. In some embodiments, the biological sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stoolWSGR Docket No. 52459-726.601 sample, a sputum sample, a lymph sample, raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof. In some embodiments, the blood sample is obtained from a subject. In some embodiments, the blood sample is collected in a blood collection tube. In some embodiments, the blood collection tube comprises a stabilizing agent for stabilizing RNAs. In some embodiments, the stabilizing agent comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid. In some embodiments, the blood sample is contacted with the sample processing buffer without removing the stabilizing agent. In some embodiments, the blood sample is contacted with the sample processing buffer directly without being subject to other processing prior to contacting the sample processing buffer. In some embodiments, the blood sample is not processed by centrifugation or a spin column prior to contacting the sample processing buffer. In some embodiments, the sample is lyophilized. In some embodiments, the sample amplification buffer is lyophilized. In some embodiments, the sample processing buffer further comprises a cucurbituril.

[0063] In some aspects, the present disclosure provides a composition for sample processing comprising: a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; a stabilizing agent comprising tetradecyl trimethyl-ammonium oxalate and / or tartaric acid, and wherein the composition is configured to stabilize an enzyme during a nucleic acid amplification, and wherein the composition is configured to reduce and / or eliminate activity of a degrading nuclease.

[0064] In some aspects, the present disclosure provides a composition for sample processing comprising: a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer; a stabilizing agent comprising tetradecyl trimethyl- ammonium oxalate and / or tartaric acid, and wherein the composition is configured to increase a rate of amplification during a nucleic acid amplification.

[0065] In some embodiments, the composition does not comprise ethanol. In some embodiments, the composition further comprises cucurbituril. In some embodiments, the cucurbituril comprises cucurbit[n]uril, wherein n is an integer of 5, 6, 7, 8, or 10. In some embodiments, the cucurbituril is cucurbit[7]uril. In some embodiments, the composition further comprises a sample. In some embodiments, the sample is a biological sample.WSGR Docket No. 52459-726.601

[0066] In some embodiments, the biological sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, a lymph sample, raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof.

[0067] In some embodiments, the blood sample is obtained from a subject. In some embodiments, the blood sample is collected in a blood collection tube. In some embodiments, the blood collection tube comprises a stabilizing agent for stabilizing RNAs. In some embodiments, the stabilizing agent comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid. In some embodiments, the blood sample is contacted with the sample processing buffer without removing the stabilizing agent. In some embodiments, the blood sample is contacted with the sample processing buffer directly without being subject to other processing prior to contacting the sample processing buffer. In some embodiments, the blood sample is not processed by centrifugation or a spin column prior to contacting the processing buffer.

[0068] In some aspects, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a lysis buffer comprising a detergent, wherein the sample comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid or wherein the sample is directly from a sample collection tube; and / or (b) contacting the sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing the sample to generate a processed sample in a mixture comprising the detergent, the solubilizer, and the cyclodextrin.

[0069] In some embodiments, the method further comprises contacting the sample with a sample amplification buffer.

[0070] In some aspects, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer, wherein the composition is configured to increase a rate of amplification during a nucleic acid amplification, and wherein the sample comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid or wherein the sample is directly from a sample collection tube.

[0071] In some embodiments, the method further comprises, prior to contacting the sample with the sample amplification buffer, contacting sample with a sample processing buffer.WSGR Docket No. 52459-726.601

[0072] In some aspects, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; (b) contacting the sample with a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin; and (c) contacting the sample with a sample stabilization buffer configured to stabilize an enzyme in a nucleic acid amplification, wherein the sample comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid or wherein the sample is directly from a sample collection tube.

[0073] In some embodiments, the sample is not processed by an RNA extraction kit. In some embodiments, the kit comprises a spin-column. In some embodiments, the kit comprises a wash pellet. In some embodiments, the method does not comprise contacting with a wash buffer. In some embodiments, the method does not comprise membrane-based extraction. In some embodiments, the method further comprises subjecting the sample to the nucleic acid amplification. In some embodiments, the nucleic acid amplification comprises polymerase chain reaction (PCR) or isothermal amplification. In some embodiments, the nucleic acid amplification comprises thermocycling the sample. In some embodiments, the nucleic acid amplification generates an amplified sample. In some embodiments, a time period from the contacting in (a) to generating the amplified sample is at most 5 minutes. In some embodiments, a processing time for the sample is a time period from the contacting in (a) to generating a processed sample prior to contacting with the amplification buffer, wherein the processing time is at most 1 min, at most 50 seconds, at most 30 seconds, or at most 20 seconds. In some embodiments, the method does not comprise heating the sample. In some embodiments, the sample is a blood sample. In some embodiments, the method further comprises obtaining the sample from a subject and collecting the sample is the sample collection tube. In some embodiments, the sample processing buffer further comprises a cucurbituril.

[0074] Additional aspects and advantages of the present disclosure will become readily apparent to those skilled in this art from the following detailed description, wherein only illustrative embodiments of the present disclosure are shown and described. As will be realized, the present disclosure is capable of other and different embodiments, and its several details are capable of modifications in various obvious respects, all without departing from the disclosure. Accordingly, the drawings and description are to be regarded as illustrative in nature, and not as restrictive.WSGR Docket No. 52459-726.601INCORPORATION BY REFERENCE

[0075] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.BRIEF DESCRIPTION OF THE DRAWINGS

[0076] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings (also “Figure” and “FIG.” herein), of which:

[0077] FIG. 1 shows an exemplary schematic depicting Sample Direct preparation method. A sample can be provided and contacted with a lysis buffer. Optionally, the sample in the lysis buffer can be incubated at room temperature for a period of time. Optionally, the sample can be incubated at a higher temperature (e.g., 95°C, 98°C, 100°C, etc.). Then a recovery buffer can be added to the sample in the lysis buffer and mixed, thereby obtaining a processed sample. The recovery buffer can be liquid or lyophilized. The processed sample can be mixed with a reaction mixture for nucleic acid amplification. The recovery buffer may be lyophilized together with the reaction mixture.

[0078] FIG. 2 shows the amplification results of different swab samples using the Sample Direct preparation. Singleplex reactions amplifying target RNA sequence of Ribonuclease P protein subunit p30 (RPP30).

[0079] FIG. 3 shows the amplification results of human nasal swab samples using Sample Direct preparation. Neisseria gonorrhoeas culture, Chlamydia trachomatis culture, and RPP30 samples were run through a triplex isothermal reaction.

[0080] FIGs. 4A-4B show the amplification results of human nasal swab samples using Sample Direct preparation. FIG. 4A depicts results of Neisseria gonorrhoeae RNA, Chlamydia trachomatis RNA, and RPP30 samples were run through a triplex isothermal reaction with 1000 IFU / CFU per reaction. FIG. 4B depicts the results of NTC Sample Direct.

[0081] FIG. 5 shows the amplification results from attenuated SARS-CoV-2 virus in NP matrix, Sample Direct preparation, and the triplex isothermal reaction (SARS-CoV-2 Spike, SARS CoV-2 NSP2, and RPP30) using DTECT chemistry, with approximately 2,000 copies / reaction.WSGR Docket No. 52459-726.601

[0082] FIG. 6 shows amplification results of synthetic Monkeypox DNA (left) and Pan Orthopox (right) in NP matrix, Sample Direct preparation, and duplex isothermal reaction (e.g., DTECT chemistry) with approximately 2,000 copies / reaction.

[0083] FIG. 7 shows the results of PCR amplification of human buccal cell samples using Sample Direct preparation. Triangles show standard curve dilution series reactions.

[0084] FIG. 8 shows results of an amplification assay of blood samples using the lysis and recovery buffers as described herein. Amplification was observed for the blood samples that were processed using the lysis and recovery buffers as described herein without any heating steps.

[0085] FIG. 9 shows results of 18s RNA amplification using differential targeted endonuclease cutting technology (DTECT) chemistry. The amplification was assessed with cucurbit[7]uril alone added, cucurbit[7]uril and gamma-cyclodextrin added, or gamma-cyclodextrin alone added. The combination of cucurbit[7]uril and gamma-cyclodextrin showed lower cycle threshold (Ct) values.

[0086] FIG. 10 shows IL1RN RNA amplification using DTECT chemistry. The amplification was assessed with cucurbit[7]uril alone added, cucurbit[7]uril and gamma-cyclodextrin added, or gamma-cyclodextrin alone added. Addition of cucurbit[7]uril alone led to lower cycle threshold (Ct) values.

[0087] FIG. 11 shows MCTP1 RNA amplification using DTECT chemistry. The amplification was assessed with cucurbit[7]uril alone added, cucurbit[7]uril and gamma-cyclodextrin added, or gamma-cyclodextrin alone added. Addition of cucurbit[7]uril alone led to lower cycle threshold (Ct) values.

[0088] FIG. 12 shows a summary of results from FIGs. 9-11.

[0089] FIG. 13 shows effects of different Tris concentrations in the recovery buffer on cycle threshold values. Concentrations of 35 mM, 52.5 mM, and 17.5 mM Tris led to lowest Ct values.

[0090] FIG. 14 shows DTECT assay performance using MMLV reverse transcriptase (MMLV RT). MMLV functioned in the DTECT assay using varying control RNA dilutions.

[0091] FIGs. 15A-15O show an example of precursor steps leading to an isothermal amplification cycle according to various embodiments described herein. FIG. 15A depicts the duplexed oligo complex binding to the target nucleic acid strand. FIG. 15B depicts endonucleolytic activity on the duplexed oligo / target complex. FIG. 15C depicts a polymerase extending off of the 3' end of the target strand. FIG. 15D depicts the polymerase displacing theWSGR Docket No. 52459-726.601 duplexed guide molecule. FIG. 15E depicts endonucleolytic activity on the oligo / extension product complex. FIG. 15F depicts a polymerase extending off the 3' end of the cut oligo and displacement of the guide. FIG. 15G depicts endonuclease activity on the newly synthesized portion complementary to the target strand. FIG. 15H depicts a polymerase extending off the 3' end of the cut site and displacement of the synthesized complement to the target strand. FIG. 151 depicts the displaced complement acting as a new target for the second complementary strand duplexed oligo complex. FIG. 15J depicts the polymerase displacing the second complementary strand duplexed guide molecule. FIG. 15K depicts the completed extension on the new guide molecule. FIG. 15L depicts endonucleolytic activity on the second complementary strand oligo / extension product complex. FIG. 15M depicts a polymerase extending off the 3’ end of the cut site of the second complementary strand of the oligo / extension product complex. FIG. 15N depicts endonucleolytic activity on the newly synthesized complementary strand of the second complementary strand guide. FIG. 150 depicts the displaced and single stranded synthesized fragments as starting material for a strand displacement amplification reaction.DETAILED DESCRIPTION OF THE INVENTIONDefinitions

[0092] While various embodiments of the invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions may occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed.

[0093] The practice of some methods disclosed herein employ, unless otherwise indicated, techniques of immunology, biochemistry, chemistry, molecular biology, microbiology, cell biology, genomics and recombinant DNA. See for example Sambrook and Green, Molecular Cloning: A Laboratory Manual, 4th Edition (2012); the series Current Protocols in Molecular Biology (F. M. Ausubel, et al. eds.); the series Methods In Enzymology (Academic Press, Inc.), PCR 2: A Practical Approach (M.J. MacPherson, B.D. Hames and G.R. Taylor eds. (1995)), Harlow and Lane, eds. (1988) Antibodies, A Laboratory Manual, and Culture of Animal Cells: A Manual of Basic Technique and Specialized Applications, 6th Edition (R.I. Freshney, ed. (2010)) (which is entirely incorporated by reference herein).

[0094] As used herein, the singular forms “a,” “an,” and “the” are intended to include the plural forms as well, unless the context clearly indicates otherwise. Furthermore, to the extent that theWSGR Docket No. 52459-726.601 terms “including,” “includes,” “having,” “has,” “with,” or variants thereof are used in either the detailed description and / or the claims, such terms are intended to be inclusive in a manner similar to the term “comprising.”

[0095] The term “about” or “approximately” means within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, i.e., the limitations of the measurement system. For example, “about” can mean within one or more than one standard deviation, per the practice in the art. Alternatively, “about” can mean a range of up to 20%, up to 15%, up to 10%, up to 5%, or up to 1% of a given value.

[0096] The term “nucleotide,” as used herein, generally refers to a base-sugar-phosphate combination. A nucleotide may comprise a synthetic nucleotide. A nucleotide may comprise a nucleotide analog. A nucleotide may comprise a synthetic nucleotide analog. Nucleotides may be monomeric units of a nucleic acid sequence (e.g., deoxyribonucleic acid (DNA) and ribonucleic acid (RNA)). The term nucleotide may include ribonucleoside triphosphates adenosine triphosphate (ATP), uridine triphosphate (UTP), cytosine triphosphate (CTP), guanosine triphosphate (GTP) and deoxyribonucleoside triphosphates (dNTPs) such as dATP, dCTP, diTP, dUTP, dGTP, dTTP, or derivatives thereof. Such derivatives may include, for example, [aS]dATP, 7-deaza-dGTP and 7-deaza-dATP, and nucleotide derivatives that confer nuclease resistance on the nucleic acid molecule containing them. Synthetic nucleotide analogs may include locked nucleic acids (LNAs), bridged nucleic acids (BNAs), fluorinated nucleic acids (also known as fluoro-modified nucleic acids), and peptide nucleic acids (PNAs). As used herein, the term “locked nucleic acid” (“LNA”), generally refers to a nucleic acid analog wherein the ribose ring is “locked” with an extra bridge connecting the 2 -oxygen atom with the 4'-carbon atom of the nucleotide such as a methylene bridge (see e.g. WO 99 / 14226, which is incorporated by reference in its entirety herein). As used herein, the term “bridged nucleic acid (BNA),” generally refers to constrained or inaccessible nucleic acid molecules which have a fixed bridge structure at the 2'- or 4'-position. As used herein, “fluorinated nucleic acids” generally refer to nucleic acids which have incorporated a fluorine atom, often at the 2'- or mposition. As used herein, the term “peptide nucleic acid (PNA),” generally refers to a nucleotide analog wherein the backbone of the analog, for example a sugar backbone in DNA, is a pseudopeptide. A PNA backbone can comprise, for example, a sequence of repeated N-(2- amino-ethyl)-glycine units. A peptide nucleic acid analog can react as DNA would react in a given environment, and can additionally bind complementary nucleic acid sequences andWSGR Docket No. 52459-726.601 various proteins. Due to the non-natural backbone, PNAs can be insensitive to endonuclease cleavage in situations where an endonuclease would cleave the equivalent DNA / RNA sequence. The term “nucleotide,” as used herein, may refer to dideoxyribonucleoside triphosphates (ddNTPs) and their derivatives. Illustrative examples of dideoxyribonucleoside triphosphates may include, but are not limited to, ddATP, ddCTP, ddGTP, ddITP, and ddTTP. A nucleotide may be unlabeled or detectably labeled, such as using moieties comprising optically detectable moieties (e.g., fluorophores). Detectable labels may include, for example, radioactive isotopes, fluorescent labels, chemiluminescent labels, bioluminescent labels and enzyme labels.

[0097] The terms “polynucleotide,” “oligonucleotide,” and “nucleic acid” are used interchangeably to generally refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof, either in single-, double-, or multistranded form. A polynucleotide may be DNA. A polynucleotide may be RNA. A polynucleotide may comprise one or more nucleotide analogs (e.g., including those with an altered backbone, sugar, or nucleobase). If present, modifications to the nucleotide structure may be imparted before or after assembly of the polymer. Some non-limiting examples of analogs include: 5-bromouracil, peptide nucleic acid, xeno nucleic acid, morpholinos, glycol nucleic acids, threose nucleic acids, dideoxynucleotides, cordycepin, 7-deaza-GTP, fluorophores (e.g., rhodamine or fluorescein linked to the sugar), thiol containing nucleotides, biotin linked nucleotides, fluorescent base analogs, CpG islands, methyl-7-guanosine, methylated nucleotides, inosine, thiouridine, pseudourdine, dihydrouridine, queuosine, wyosine, PNAs, and LNAs.

[0098] As used herein, the term “restriction endonuclease,” “restriction enzyme,” or grammatical equivalents thereof generally refers to an enzyme that originates in bacterial host defense and is understood to recognize a specific sequence on an incoming viral DNA and cleave the DNA either at the recognition sequence or at a distinct sequence site. One group of restriction endonucleases are identified as Type IIS. This group can recognize asymmetric DNA sequences and cleaves the DNA at a site outside the cleavage site that is at a defined distance from the recognition site. In some cases, type IIS restriction endonucleases cleave DNA between 1 and 20 nucleotides from the relevant recognition site.

[0099] As used herein, the term “restriction endonuclease recognition sequence” generally refers to a location on a nucleic acid molecule (e.g., DNA molecule) containing specific sequences of nucleotides, which are recognized by various restriction enzymes. These sequences canWSGR Docket No. 52459-726.601 comprise from 4-8 base pairs to 12-40 base pairs in length. These sites can be palindromic sequences.

[0100] As used herein, the term “sample” refers to a substance (e.g., solid or liquid) that contains a target nucleic acid sequence to be amplified. The target nucleic acid may be DNA. The target nucleic acid molecule may be RNA. As used herein, a “processed sample” refers to a sample that has been contacted by a lysis buffer and / or a recovery buffer of the present disclosure.

[0101] As used herein, the term “template” generally refers to a portion of a target RNA or DNA of a sample that is amplified by a DNA polymerase to produce one or more amplified nucleic acid products.

[0102] As used herein, “amplified product”, “amplified nucleic acid product”, or “amplicon” generally refers to the end product resulting from a nucleic acid method, such as PCR or isothermal amplification.

[0103] As used herein, the term “polymerase” generally refers to an enzyme that produces a complementary replicate of a nucleic acid molecule using the nucleic acid as a template strand. DNA polymerases bind to the template strand and then move down the template strand adding nucleotides to the free hydroxyl group at the 3' end of a growing chain of nucleic acid. DNA polymerases synthesize complementary DNA molecules from DNA (e.g. DNA-dependent DNA polymerases) or RNA templates (e.g. RNA-dependent DNA polymerases or reverse transcriptases) and RNA polymerases synthesize RNA molecules from DNA templates (e.g. DNA-dependent RNA polymerases which participate in transcription). DNA polymerases generally use a short, preexisting RNA or DNA strand, called a primer, to begin chain growth. Some DNA polymerases replicate single-stranded templates, while other DNA polymerases displace the strand upstream of the site where they add bases to a chain.

[0104] As used herein, the term “strand displacing,” when used in reference to a polymerase, generally refers to an activity that removes a complementary strand from base-pairing with a template strand being read by the polymerase. Example polymerases having strand displacing activity include the large fragment of Bacillus stearothermophilus polymerase, exo-Klenow polymerase, B st 2.0 polymerase, B st 3.0 polymerase, SD DNA polymerase, phi29 DNA polymerase, and sequencing-grade T7 exo-polymerase.

[0105] As used herein, “primer”, or “primer sequence” generally refers to a linear oligonucleotide that is complementary to and anneals to a target sequence. The lower limit on primer length is determined by ability to hybridize since very short primers (e.g., less than 5WSGR Docket No. 52459-726.601 nucleotides) do not form thermodynamically stable duplexes under most hybridization conditions. Primers may vary in length from 4 to 50 nucleotides. In some embodiments, the primer is between about 10 and 20 nucleotides in length. In some embodiments, the primer can be more than about 100 nucleotides in length. In some embodiments, the primer can be an oligonucleotide that can hybridize with a target nucleic acid sequence. In some embodiments, the primer can be a probe. In some embodiments, the primer can comprise a guide oligonucleotide. For example, the primer (e.g., the guide oligonucleotide) can be an oligonucleotide comprising a target binding region that hybridizes to a target polynucleotide sequence and a non-target binding region that does not hybridize to a target sequence. In some embodiments, the non-target binding region of the primer can comprise a palindromic sequence. In some embodiments, the palindromic sequence may permit recruitment of binding of a restriction enzyme to process the target sequence.

[0106] As used herein, the terms “amplify,” “amplifies,” “amplified,” “amplification,” and “amplicon” generally refer to any method for replicating a nucleic acid. The replication can be conducted with the use of a primer-dependent polymerase. The replication can be enzyme-free amplification. In some cases, amplifying or replicating a target nuclei acid strand also comprises replicating or amplifying a complementary strand of the target nucleic acid strand. Amplified products can be subjected to subsequence analyses, including but not limited to melting curve analysis, nucleotide sequencing, single-strand conformation polymorphism assay, allele-specific oligonucleotide hybridization, Southern blot analysis, and restriction endonuclease digestion.

[0107] The terms “hybridizes,” and “annealing,” as used herein, generally refer to a reaction in which one or more polynucleotides interact to form a complex that is stabilized via hydrogen bonding between the bases of the nucleotide residues. The hydrogen bonding may occur by Watson Crick base pairing, Hoogstein binding, or in any other sequence sensitive or specific manner. The complex may comprise two strands forming a duplex structure, three or more strands forming a multi stranded complex, a single self-hybridizing strand, or any combination of these. A hybridization reaction may constitute a step in a more extensive process, such as the initiation of a PCR, or the enzymatic cleavage of a polynucleotide by a ribozyme. A first sequence that can be stabilized via hydrogen bonding with the bases of the nucleotide residues of a second sequence can generally be “hybridizable” to the second sequence. In such a case, the second sequence can also be the to be hybridizable to the first sequence.WSGR Docket No. 52459-726.601

[0108] The terms “complement,” “complements,” “complementary,” and “complementarity,” as used herein, generally refer to a sequence that is fully complementary to and hybridizable to the given sequence. In some cases, a first sequence that is hybridizable to a second sequence or set of second sequences is specifically or selectively hybridizable to the second sequence or set of second sequences, such that hybridization to the second sequence or set of second sequences is used. Hybridizable sequences can share a degree of sequence complementarity over all or a portion of their respective lengths, such as between 25%-100% complementarity, including at least about 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, and 100% sequence complementarity.Overview

[0109] The present application provides compositions of sample processing buffers, sample stabilization buffer, and amplification reaction buffers, kits containing one or more of the buffers described herein, and method of using the compositions. The sample processing buffers, sample stabilization buffer, and amplification reaction buffers can be mixed directly at the appropriate steps without the need of washing steps or removing the buffers from any prior steps. The compositions, methods, and kits described herein can be used as part of a framework system to enhance research and development processes of samples (e.g., biological samples). The compositions, methods, and kits described herein, and uses thereof, can be designed to be flexible and adaptable, reducing the time necessary to develop products.

[0110] The buffer compositions described herein, e.g., lysis buffers, recovery buffers, amplifications and / or stabilization buffers, can be versatile and can function with various component concentrations.

[0111] The compositions and methods described herein can be used to process various samples and can function in the presence of any inhibitors that may be present in a sample.

[0112] Compositions described herein can be part of an amplification buffer system, a sample processing buffer system, a stabilization buffer system, or any combination thereof. The amplification buffer (e.g., core amplification buffer) can comprise enzymes, primers and / or probes (e.g., guide oligonucleotides), reverse transcriptase primers, molecular beacons, dNTPs, or any combination thereof. The sample processing buffer (e.g., Sample Direct) can comprise reagents of lysis and / or recovery buffers described herein. The sample processing buffer can comprise salts and / or buffers which may be adjusted to optimize amplification reactions (e.g., PCR and / or isothermal amplification). The stabilization buffer can comprise cyclodextrin,WSGR Docket No. 52459-726.601 protein stabilizers, cake structure modifiers (Tc, Tg, Tg’), salts, buffers, or any combination thereof. Cake structure modifiers can comprise reagents that modify the glass transition temperature (Tg), the glass transition temperature of the maximally freeze concentrated master mix solute prior to being dried (Tg’), the onset crystallization temperature (Tc), or any combination thereof. One or more cake structure modifiers may increase a critical collapse temperature of a composition described herein (e.g., a sample stabilization buffer). In some embodiments, one or more cake structure modifiers may enable a more efficient (e.g., warmer) drying cycle for the composition described herein (e.g., the sample stabilization buffer). The cake structure modifiers may improve structural properties of the dried composition (e.g., dried cake). The enhanced structural properties may make the dried composition (e.g., dried cake) more resistant to crushing, fracture, cracking, or any combination thereof. In some cases, glass transition temperatures can vary, for example from about 140 °C to 370 °C. The reagents of the stabilization buffer may be optimized for freeze drying.

[0113] Screening assays in a representative sample matrix can decrease risks of downstream sample-assay integration. Consistent drying cycles can be available for immediate research and development use. The benefits of the compositions, methods, and kits described herein can include (i) reducing the time to develop and integrate assays into a commercially viable shelfstable formulation; (ii) screening and optimizing assays in a representative sample matrix (e.g., matrix-based screening, compositions of the total sample types), (iii) a baked-in excipient that may be lyophilized with a compatible freeze-drying cycle, and (iv) eliminating the distinction between chemistry (e.g., chemical reagents) intended to be run immediately (for example research and development experiments) and chemistry which are intended to be freeze-dried. The unification of the compositions, methods, and kits described herein provide for greater efficiency in sample processing, stabilization, and amplification.

[0114] The methods using the compositions described herein can be referred to as “Sample Direct” preparation methods. The methods provided herein can process a sample quickly (e.g., at most about 5 minutes, at most about 4 minutes, at most about 3 minutes, at most about 2 minutes, at most about 1 minute, at most about 45 seconds, at most about 30 seconds, at most about 20 seconds, or less) and improve amplification reaction performance.Compositions for Sample Processing, Stabilization, and AmplificationWSGR Docket No. 52459-726.601

[0115] The present disclosure provides compositions and methods for processing a sample comprising a target nucleic acid molecule for nucleic acid amplification. The target nucleic acid molecule may be DNA and / or RNA.

[0116] In some aspects, the present disclosure provides compositions for sample processing for a nucleic acid amplification method. In some embodiments, the composition comprises a detergent, a solubilizer, and a cyclodextrin. Without wishing to be bound by theory, the composition may be configured to stabilize an enzyme during the nucleic acid amplification. The composition may also assist in reducing the activity of a degrading nuclease during the nucleic acid amplification. The composition may eliminate the activity of a degrading nuclease during the nucleic acid amplification. The composition may degrade or inactivate the function of a nuclease prior to the nucleic acid amplification. The composition may be configured to lyse cell walls and / or nuclear membranes.

[0117] In some embodiments, the enzyme stabilized by the composition provided herein is a polymerase, an endonuclease, a reverse transcriptase, a ligase, a helicase, a recombinase, or any combination thereof.

[0118] In some embodiments, the nuclease is a ribonuclease. In some embodiments, the ribonuclease comprises an endoribonuclease or an exoribonuclease. In some embodiments, the endoribonuclease includes, but is not limited to, RNAase A, RNAase H, RNAase III, RNAase L, RNAase P, RNAase PhyM, RNAase Tl, RNAase T2, RNAase U2, RNAase V, RNAase E, and RNAase G. In some embodiments, the exoribonuclease includes, but is not limited to, RNAase PH, RNAase R, RNAase D, RNAase T, oligoribonuclease, exoribonuclease I, exoribonuclease II, and polynucleotide phosphorylase (e.g., PNPase).

[0119] In some embodiments, the detergent is sodium dodecyl sulfate (SDS). In some embodiments, the detergent comprises sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the detergent is an ionic detergent. In some embodiments, the detergent is a non-ionic detergent. In some embodiments, the detergent is part of a lysis buffer. A lysis buffer is capable of lysing cells yet leaving nucleic acids intact (e.g., not denaturing a nucleic acid chain to the extent that the chain is disrupted to individual nucleic acids). In some embodiments, the lysis buffer is capable of handling challenging solid and liquid sample types.

[0120] In some embodiments, the detergent is present at a final concentration when mixed with the sample to be processed in the lysis buffer. The detergent may be present at a final concentration that is effective for lysing cells in the mixture in the presence of the sample. TheWSGR Docket No. 52459-726.601 concentration of any agent described herein when mixed with a sample to be processed can be referred to as final concentration. In some embodiments, the concentration (e.g., final concentration) of the detergent in the mixture in the presence of the sample is at least about 0.05% w / v (where w / v refers to g of solute / 100 mL of solution), at least about 0.1% w / v, at least about 0.15% w / v, at least about 0.2% w / v, at least about 0.25% w / v, at least about 0.3% w / v, at least about 0.35% w / v, at least about 0.4% w / v, at least about 0.45% w / v, at least about 0.5% w / v, at least about 0.55% w / v, at least about 0.6% w / v, at least about 0.65% w / v, at least about 0.7% w / v, at least about 0.75% w / v, at least about 0.8% w / v, at least about 0.85% w / v, at least about 0.9% w / v, at least about 0.95% w / v, at least about 1.0% w / v, at least about 2.0% w / v, at least about 3.0% w / v, at least about 4.0% w / v, at least about 5.0% w / v, at least about 6.0% w / v, at least about 7.0% w / v, at least about 8.0% w / v, at least about 9.0% w / v, or at least about 10.0% w / v.

[0121] In some embodiments, the concentration (e.g., final concentration) of the detergent in the mixture in the presence of the sample is at most about 10.0% w / v, at most about 9.0% w / v, at most about 8.0% w / v, at most about 7.0% w / v, at most about 6.0% w / v, at most about 5.0% w / v, at most about 4.0% w / v, at most about 3.0% w / v, at most about 2.0% w / v, at most about 1.0% w / v, at most about 0.95% w / v, at most about 0.9% w / v, at most about 0.85% w / v, at most about 0.8% w / v, at most about 0.75% w / v, at most about 0.7% w / v, at most about 0.65% w / v, at most about 0.6% w / v, at most about 0.55% w / v, at most about 0.5% w / v, at most about 0.45% w / v, at most about 0.4% w / v, at most about 0.35% w / v, at most about 0.3% w / v, at most about 0.25% w / v, at most about 0.2% w / v, at most about 0.15% w / v, at most about 0.1% w / v, or at most about 0.05% w / v.

[0122] In some embodiments, the concentration (e.g., final concentration) of the detergent in the mixture in the presence of the sample is about 0.1% w / v to about 2% w / v. In some embodiments, the concentration (e.g., final concentration) of the detergent in the mixture in the presence of the sample is about 0.1% w / v to about 0.2% w / v, about 0.1% w / v to about 0.3% w / v, about 0.1% w / v to about 0.4% w / v, about 0.1% w / v to about 0.5% w / v, about 0.1% w / v to about 0.6% w / v, about 0.1% w / v to about 0.7% w / v, about 0.1% w / v to about 0.8% w / v, about 0.1% w / v to about 0.9% w / v, about 0.1% w / v to about 1% w / v, about 0.1% w / v to about 1.5% w / v, about 0.1% w / v to about 2% w / v, about 0.2% w / v to about 0.3% w / v, about 0.2% w / v to about 0.4% w / v, about 0.2% w / v to about 0.5% w / v, about 0.2% w / v to about 0.6% w / v, about 0.2% w / v to about 0.7% w / v, about 0.2% w / v to about 0.8% w / v, about 0.2% w / v to about 0.9% w / v, about 0.2% w / v to about 1% w / v, about 0.2% w / v to about 1.5% w / v, about 0.2% w / v toWSGR Docket No. 52459-726.601 about 2% w / v, about 0.3% w / v to about 0.4% w / v, about 0.3% w / v to about 0.5% w / v, about 0.3% w / v to about 0.6% w / v, about 0.3% w / v to about 0.7% w / v, about 0.3% w / v to about 0.8% w / v, about 0.3% w / v to about 0.9% w / v, about 0.3% w / v to about 1% w / v, about 0.3% w / v to about 1.5% w / v, about 0.3% w / v to about 2% w / v, about 0.4% w / v to about 0.5% w / v, about 0.4% w / v to about 0.6% w / v, about 0.4% w / v to about 0.7% w / v, about 0.4% w / v to about 0.8% w / v, about 0.4% w / v to about 0.9% w / v, about 0.4% w / v to about 1% w / v, about 0.4% w / v to about 1.5% w / v, about 0.4% w / v to about 2% w / v, about 0.5% w / v to about 0.6% w / v, about 0.5% w / v to about 0.7% w / v, about 0.5% w / v to about 0.8% w / v, about 0.5% w / v to about 0.9% w / v, about 0.5% w / v to about 1% w / v, about 0.5% w / v to about 1.5% w / v, about 0.5% w / v to about 2% w / v, about 0.6% w / v to about 0.7% w / v, about 0.6% w / v to about 0.8% w / v, about 0.6% w / v to about 0.9% w / v, about 0.6% w / v to about 1% w / v, about 0.6% w / v to about 1.5% w / v, about 0.6% w / v to about 2% w / v, about 0.7% w / v to about 0.8% w / v, about 0.7% w / v to about 0.9% w / v, about 0.7% w / v to about 1% w / v, about 0.7% w / v to about 1.5% w / v, about 0.7% w / v to about 2% w / v, about 0.8% w / v to about 0.9% w / v, about 0.8% w / v to about 1% w / v, about 0.8% w / v to about 1.5% w / v, about 0.8% w / v to about 2% w / v, about 0.9% w / v to about 1% w / v, about 0.9% w / v to about 1.5% w / v, about 0.9% w / v to about 2% w / v, about 1% w / v to about 1.5% w / v, about 1% w / v to about 2% w / v, or about 1.5% w / v to about 2% w / v.

[0123] In some embodiments, the lysis buffer further comprises additional agents including, but not limited to, egtazic acid (EGTA), ethylenediaminetetraacetic acid (EDTA), tris(2- carboxyethyl)phosphine (TCEP), and / or tris(hydroxymethyl)aminomethane (e.g., Tris). In some embodiments, the lysis buffer comprises SDS, egtazic acid (EGTA), ethylenediaminetetraacetic acid (EDTA), tris(2-carboxyethyl)phosphine (TCEP), and / or tri s(hydroxymethyl)aminom ethane (c.g, Tris). In some embodiments, the lysis buffer further comprises lysis buffer comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2- carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

[0124] In some embodiments, the lysis buffer comprises a chelating agent. In some embodiments, the lysis buffer comprises 1, 2, 3, 4, or more chelating agents. In some embodiments, the chelating agent comprises is deferiprone, ethylenediamine, 1,10-WSGR Docket No. 52459-726.601Phenanthroline, oxalic acid, pentetic acid, deferasirox, deferoxamine, deferoxamine mesylate, or N,N,N',N'-tetrakis(2-pyridinylmethyl)-l ,2-ethanediamine (TPEN).

[0125] In some embodiments, the lysis buffer comprises a reducing agent. In some embodiments, the lysis buffer comprises 1, 2, 3, 4, 5, or more reducing agents. In some embodiments, the reducing agent comprises oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

[0126] In some embodiments, EGTA is present at a final concentration in the lysis buffer effective for binding calcium, magnesium, and / or other ions. In some embodiments, the concentration (e.g., final concentration) of EGTA in the mixture in the presence of the sample is at least about 0.1 mM, at least about 0.25 mM, at least about 0.5 mM, at least about 0.75 mM, at least about 1.0 mM, at least about 1.5 mM, at least about 2.0 mM, at least about 2.5 mM, at least about 3.0 mM, at least about 3.5 mM, at least about 4.0 mM, at least about 4.5 mM, at least about 5.0 mM, at least about 5.5 mM, at least about 6.0 mM, at least about 6.5 mM, at least about 7.0 mM, at least about 7.5 mM, at least about 8.0 mM, at least about 8.5 mM, at least about 9.0 mM, at least about 9.5 mM, at least about 10.0 mM, at least about 11.0 mM, at least about 12.0 mM, at least about 13.0 mM, at least about 14.0 mM, at least about 15.0 mM, at least about 20.0 mM, at least about 25.0 mM, at least about 30.0 mM, at least about 35.0 mM, at least about 40.0 mM, at least about 45.0 mM, or at least about 50.0 mM.

[0127] In some embodiments, the concentration (e.g., final concentration) of EGTA in the mixture in the presence of the sample is at most about 50.0 mM, at most about 45.0 mM, at most about 40.0 mM, at most about 35.0 mM, at most about 30.0 mM, at most about 25.0 mM, at most about 20.0 mM, at most about 15.0 mM, at most about 10.0 mM, at most about 9.5 mM, at most about 9.0 mM, at most about 8.5 mM, at most about 8.0 mM, at most about 7.5 mM, at most about 7.0 mM, at most about 6.5 mM, at most about 6.0 mM, at most about 5.5 mM, at most about 5.0 mM, at most about 4.5 mM, at most about 4.0 mM, at most about 3.5 mM, at most about 3.0 mM, at most about 2.5 mM, at most about 2.0 mM, at most about 1.5 mM, at most about 1.0 mM, at most about 0.75 mM, at most about 0.5 mM, at most about 0.25 mM, or at most about 0.1 mM.

[0128] In some embodiments, the concentration (e.g., final concentration) of EGTA in the mixture in the presence of the sample is about 0.5 mM to about 20 mM. In some embodiments, the concentration (e.g., final concentration) of EGTA in the mixture in the presence of the sample is at most about 20 mM. In some embodiments, the concentration (e.g., finalWSGR Docket No. 52459-726.601 concentration) of EGTA in the mixture in the presence of the sample is about 0.5 mM to about 1 mM, about 0.5 mM to about 2 mM, about 0.5 mM to about 3 mM, about 0.5 mM to about 4 mM, about 0.5 mM to about 5 mM, about 0.5 mM to about 6 mM, about 0.5 mM to about 8 mM, about 0.5 mM to about 10 mM, about 0.5 mM to about 12 mM, about 0.5 mM to about 15 mM, about 0.5 mM to about 20 mM, about 1 mM to about 2 mM, about 1 mM to about 3 mM, about 1 mM to about 4 mM, about 1 mM to about 5 mM, about 1 mM to about 6 mM, about 1 mM to about 8 mM, about 1 mM to about 10 mM, about 1 mM to about 12 mM, about 1 mM to about 15 mM, about 1 mM to about 20 mM, about 2 mM to about 3 mM, about 2 mM to about 4 mM, about 2 mM to about 5 mM, about 2 mM to about 6 mM, about 2 mM to about 8 mM, about 2 mM to about 10 mM, about 2 mM to about 12 mM, about 2 mM to about 15 mM, about 2 mM to about 20 mM, about 3 mM to about 4 mM, about 3 mM to about 5 mM, about 3 mM to about 6 mM, about 3 mM to about 8 mM, about 3 mM to about 10 mM, about 3 mM to about 12 mM, about 3 mM to about 15 mM, about 3 mM to about 20 mM, about 4 mM to about 5 mM, about 4 mM to about 6 mM, about 4 mM to about 8 mM, about 4 mM to about 10 mM, about 4 mM to about 12 mM, about 4 mM to about 15 mM, about 4 mM to about 20 mM, about 5 mM to about 6 mM, about 5 mM to about 8 mM, about 5 mM to about 10 mM, about 5 mM to about 12 mM, about 5 mM to about 15 mM, about 5 mM to about 20 mM, about 6 mM to about 8 mM, about 6 mM to about 10 mM, about 6 mM to about 12 mM, about 6 mM to about 15 mM, about 6 mM to about 20 mM, about 8 mM to about 10 mM, about 8 mM to about 12 mM, about 8 mM to about 15 mM, about 8 mM to about 20 mM, about 10 mM to about 12 mM, about 10 mM to about 15 mM, about 10 mM to about 20 mM, about 12 mM to about 15 mM, about 12 mM to about 20 mM, or about 15 mM to about 20 mM.

[0129] In some embodiments, EDTA is present at a final concentration in the lysis buffer effective for binding magnesium, calcium, and / or other ions. In some embodiments, the concentration (e.g., final concentration) of EDTA in the mixture in the presence of the sample is at least about 0.1 mM, at least about 0.2, at least about 0.3 mM, at least about 0.4 mM, at least about 0.5 mM, at least about 0.6 mM, at least about 0.7 mM, at least about 0.8 mM, at least about 0.9 mM, at least about 1.0 mM, at least about 1.1 mM, at least about 1.2 mM, at least about 1.3 mM, at least about 1.4 mM, at least about 1.5 mM, at least about 1.6 mM, at least about 1.7 mM, at least about 1.8 mM, at least about 1.9 mM, at least about 2.0 mM, at least about 3.0 mM, at least about 4.0 mM, at least about 5.0 mM, at least about 6.0 mM, at least about 7.0 mM, at least about 8.0 mM, at least about 9.0 mM, at least about 10.0 mM, at least about 20.0 mM, at least about 30.0 mM, at least about 40.0 mM, or at least about 50.0 mM.WSGR Docket No. 52459-726.601

[0130] In some embodiments, the concentration (e.g., final concentration) of EDTA in the mixture in the presence of the sample is at most about 50.0 mM, at most about 40.0 mM, at most about 30.0 mM, at most about 20.0 mM, at most about 10.0 mM, at most about 9.0 mM, at most about 8.0 mM, at most about 7.0 mM, at most about 6.0 mM, at most about 5.0 mM, at most about 4.0 mM, at most about 3.0 mM, at most about 2.0 mM, at most about 1.9 mM, at most about 1.8 mM, at most about 1.7 mM, at most about 1.6 mM, at most about 1.5 mM, at most about 1.4 mM, at most about 1.3 mM, at most about 1.2 mM, at most about 1.1 mM, at most about 1.0 mM, at most about 0.9 mM, at most about 0.8 mM, at most about 0.7 mM, at most about 0.6 mM, at most about 0.5 mM, at most about 0.4 mM, at most about 0.3 mM, at most about 0.2 mM, or at most about 0.1 mM.

[0131] In some embodiments, the concentration (e.g., final concentration) of EDTA in the mixture in the presence of the sample is about 0.1 mM to about 25 mM. In some embodiments, the concentration (e.g., final concentration) of EDTA in the mixture in the presence of the sample is at most about 25 mM. In some embodiments, the concentration (e.g., final concentration) of EDTA in the mixture in the presence of the sample is about 0.1 mM to about 0.25 mM, about 0.1 mM to about 0.5 mM, about 0.1 mM to about 0.75 mM, about 0.1 mM to about 1 mM, about 0.1 mM to about 1.5 mM, about 0.1 mM to about 2 mM, about 0.1 mM to about 2.5 mM, about 0.1 mM to about 3 mM, about 0.1 mM to about 5 mM, about 0.1 mM to about 10 mM, about 0.1 mM to about 25 mM, about 0.25 mM to about 0.5 mM, about 0.25 mM to about 0.75 mM, about 0.25 mM to about 1 mM, about 0.25 mM to about 1.5 mM, about 0.25 mM to about 2 mM, about 0.25 mM to about 2.5 mM, about 0.25 mM to about 3 mM, about 0.25 mM to about 5 mM, about 0.25 mM to about 10 mM, about 0.25 mM to about 25 mM, about 0.5 mM to about 0.75 mM, about 0.5 mM to about 1 mM, about 0.5 mM to about 1.5 mM, about 0.5 mM to about 2 mM, about 0.5 mM to about 2.5 mM, about 0.5 mM to about 3 mM, about 0.5 mM to about 5 mM, about 0.5 mM to about 10 mM, about 0.5 mM to about 25 mM, about 0.75 mM to about 1 mM, about 0.75 mM to about 1.5 mM, about 0.75 mM to about 2 mM, about 0.75 mM to about 2.5 mM, about 0.75 mM to about 3 mM, about 0.75 mM to about 5 mM, about 0.75 mM to about 10 mM, about 0.75 mM to about 25 mM, about 1 mM to about 1.5 mM, about 1 mM to about 2 mM, about 1 mM to about 2.5 mM, about 1 mM to about 3 mM, about 1 mM to about 5 mM, about 1 mM to about 10 mM, about 1 mM to about 25 mM, about 1.5 mM to about 2 mM, about 1.5 mM to about 2.5 mM, about 1.5 mM to about 3 mM, about 1.5 mM to about 5 mM, about 1.5 mM to about 10 mM, about 1.5 mM to about 25 mM, about 2 mM to about 2.5 mM, about 2 mM to about 3 mM, about 2 mM to about 5 mM, about 2WSGR Docket No. 52459-726.601 mM to about 10 mM, about 2 mM to about 25 mM, about 2.5 mM to about 3 mM, about 2.5 mM to about 5 mM, about 2.5 mM to about 10 mM, about 2.5 mM to about 25 mM, about 3 mM to about 5 mM, about 3 mM to about 10 mM, about 3 mM to about 25 mM, about 5 mM to about 10 mM, about 5 mM to about 25 mM, or about 10 mM to about 25 mM.

[0132] In some embodiments, the reducing agent comprises tris(2-carboxyethyl)phosphine (TCEP). In some embodiments, the concentration e.g., final concentration) of TCEP in the mixture in the presence of the sample is at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, or at least about 100 mM.

[0133] In some embodiments, the concentration (e.g., final concentration) of TCEP in the mixture in the presence of the sample is at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, or at most about 1 mM.

[0134] In some embodiments, the concentration (e.g., final concentration) of TCEP in the mixture in the presence of the sample is about 0.5 mM to about 75 mM. In some embodiments, the concentration (e.g., final concentration) of TCEP in the mixture in the presence of the sample is about 0.5 mM to about 1 mM, about 0.5 mM to about 2 mM, about 0.5 mM to about 3 mM, about 0.5 mM to about 5 mM, about 0.5 mM to about 10 mM, about 0.5 mM to about 12 mM, about 0.5 mM to about 15 mM, about 0.5 mM to about 20 mM, about 0.5 mM to about 25 mM, about 0.5 mM to about 50 mM, about 0.5 mM to about 75 mM, about 1 mM to about 2 mM, about 1 mM to about 3 mM, about 1 mM to about 5 mM, about 1 mM to about 10 mM, about 1 mM to about 12 mM, about 1 mM to about 15 mM, about 1 mM to about 20 mM, about 1 mM to about 25 mM, about 1 mM to about 50 mM, about 1 mM to about 75 mM, about 2 mMWSGR Docket No. 52459-726.601 to about 3 mM, about 2 mM to about 5 mM, about 2 mM to about 10 mM, about 2 mM to about 12 mM, about 2 mM to about 15 mM, about 2 mM to about 20 mM, about 2 mM to about 25 mM, about 2 mM to about 50 mM, about 2 mM to about 75 mM, about 3 mM to about 5 mM, about 3 mM to about 10 mM, about 3 mM to about 12 mM, about 3 mM to about 15 mM, about 3 mM to about 20 mM, about 3 mM to about 25 mM, about 3 mM to about 50 mM, about 3 mM to about 75 mM, about 5 mM to about 10 mM, about 5 mM to about 12 mM, about 5 mM to about 15 mM, about 5 mM to about 20 mM, about 5 mM to about 25 mM, about 5 mM to about 50 mM, about 5 mM to about 75 mM, about 10 mM to about 12 mM, about 10 mM to about 15 mM, about 10 mM to about 20 mM, about 10 mM to about 25 mM, about 10 mM to about 50 mM, about 10 mM to about 75 mM, about 12 mM to about 15 mM, about 12 mM to about 20 mM, about 12 mM to about 25 mM, about 12 mM to about 50 mM, about 12 mM to about 75 mM, about 15 mM to about 20 mM, about 15 mM to about 25 mM, about 15 mM to about 50 mM, about 15 mM to about 75 mM, about 20 mM to about 25 mM, about 20 mM to about 50 mM, about 20 mM to about 75 mM, about 25 mM to about 50 mM, about 25 mM to about 75 mM, or about 50 mM to about 75 mM.

[0135] In some embodiments, the reducing agent comprises a Tris. In some embodiments, the concentration (e.g., final concentration) of a Tris in the mixture in the presence of the sample is at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, or at least about 100 mM.

[0136] In some embodiments, the concentration (e.g., final concentration) of a Tris in the mixture in the presence of the sample is at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at mostWSGR Docket No. 52459-726.601 about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, or at most about 1 mM.

[0137] In some embodiments, a Tris is present at a concentration e.g., final concentration) in the mixture in the presence of the sample of at least about 0.5 mM to about 75 mM. In some embodiments, the concentration (e.g., final concentration) of Tris in the mixture in the presence of the sample is about 0.5 mM to about 1 mM, about 0.5 mM to about 2 mM, about 0.5 mM to about 3 mM, about 0.5 mM to about 5 mM, about 0.5 mM to about 10 mM, about 0.5 mM to about 12 mM, about 0.5 mM to about 15 mM, about 0.5 mM to about 20 mM, about 0.5 mM to about 25 mM, about 0.5 mM to about 50 mM, about 0.5 mM to about 75 mM, about 1 mM to about 2 mM, about 1 mM to about 3 mM, about 1 mM to about 5 mM, about 1 mM to about 10 mM, about 1 mM to about 12 mM, about 1 mM to about 15 mM, about 1 mM to about 20 mM, about 1 mM to about 25 mM, about 1 mM to about 50 mM, about 1 mM to about 75 mM, about2 mM to about 3 mM, about 2 mM to about 5 mM, about 2 mM to about 10 mM, about 2 mM to about 12 mM, about 2 mM to about 15 mM, about 2 mM to about 20 mM, about 2 mM to about 25 mM, about 2 mM to about 50 mM, about 2 mM to about 75 mM, about 3 mM to about 5 mM, about 3 mM to about 10 mM, about 3 mM to about 12 mM, about 3 mM to about 15 mM, about 3 mM to about 20 mM, about 3 mM to about 25 mM, about 3 mM to about 50 mM, about3 mM to about 75 mM, about 5 mM to about 10 mM, about 5 mM to about 12 mM, about 5 mM to about 15 mM, about 5 mM to about 20 mM, about 5 mM to about 25 mM, about 5 mM to about 50 mM, about 5 mM to about 75 mM, about 10 mM to about 12 mM, about 10 mM to about 15 mM, about 10 mM to about 20 mM, about 10 mM to about 25 mM, about 10 mM to about 50 mM, about 10 mM to about 75 mM, about 12 mM to about 15 mM, about 12 mM to about 20 mM, about 12 mM to about 25 mM, about 12 mM to about 50 mM, about 12 mM to about 75 mM, about 15 mM to about 20 mM, about 15 mM to about 25 mM, about 15 mM to about 50 mM, about 15 mM to about 75 mM, about 20 mM to about 25 mM, about 20 mM to about 50 mM, about 20 mM to about 75 mM, about 25 mM to about 50 mM, about 25 mM to about 75 mM, or about 50 mM to about 75 mM.

[0138] In some embodiments, the lysis buffer can comprise SDS, EGTA, EDTA, TCEP and / or Tris. The lysis buffer can comprise SDS with a final concentration of about 0.01% w / v to 0.4% w / v in the presence of the sample; EGTA with a final concentration of about 0.1 mM to 3 mM in the presence of the sample; EDTA with a final concentration of about 0.01 mM to 1 mM in the presence of the sample; TCEP with a final concentration of about 1.0 mM to 4.0 mM in the presence of the sample; and Tris with a final concentration of about 1.0 mM to 4.5 mM in theWSGR Docket No. 52459-726.601 presence of the sample. For example, the lysis buffer used can comprise 0.2% SDS, 2 mM EGTA, 0.5 mM EDTA, 1 mM TCEP, and 1 mM Tris final concentration for each component in the presence of the sample.

[0139] In some embodiments, the lysis buffer has a pH value sufficient to lyse a desired cell. In some embodiments, the lysis buffer has a pH of at least about 1, at least about 2, at least about 3, at least about 4, at least about 4.5, at least about 5, at least about 5.5, at least about 6, at least about 6.5, at least about 7, at least about 8, or at least about 9. In some embodiments, the lysis buffer has a pH value of at most about 9, at most about 8, at most about 7, at most about 6.5, at most about 6, at most about 5.5, at most about 5, at most about 4.5, at most about 4, at most about 3, at most about 2, or at most about 1.

[0140] In some embodiments, the lysis buffer has a pH of about 1 to about 10. In some embodiments, the lysis buffer has a pH of at most about 10. In some embodiments, the lysis buffer has a pH of about 1 to about 2, about 1 to about 3, about 1 to about 4, about 1 to about 5, about 1 to about 5.5, about 1 to about 6, about 1 to about 6.5, about 1 to about 7, about 1 to about 8, about 1 to about 9, about 1 to about 10, about 2 to about 3, about 2 to about 4, about 2 to about 5, about 2 to about 5.5, about 2 to about 6, about 2 to about 6.5, about 2 to about 7, about 2 to about 8, about 2 to about 9, about 2 to about 10, about 3 to about 4, about 3 to about 5, about 3 to about 5.5, about 3 to about 6, about 3 to about 6.5, about 3 to about 7, about 3 to about 8, about 3 to about 9, about 3 to about 10, about 4 to about 5, about 4 to about 5.5, about 4 to about 6, about 4 to about 6.5, about 4 to about 7, about 4 to about 8, about 4 to about 9, about 4 to about 10, about 5 to about 5.5, about 5 to about 6, about 5 to about 6.5, about 5 to about 7, about 5 to about 8, about 5 to about 9, about 5 to about 10, about 5.5 to about 6, about 5.5 to about 6.5, about 5.5 to about 7, about 5.5 to about 8, about 5.5 to about 9, about 5.5 to about 10, about 6 to about 6.5, about 6 to about 7, about 6 to about 8, about 6 to about 9, about 6 to about 10, about 6.5 to about 7, about 6.5 to about 8, about 6.5 to about 9, about 6.5 to about 10, about 7 to about 8, about 7 to about 9, about 7 to about 10, about 8 to about 9, about 8 to about 10, or about 9 to about 10. In some embodiments, the lysis buffer has a pH of about 8. In some embodiments, the lysis buffer has a pH of about 7. In some embodiments, the lysis buffer has a pH of about 6.

[0141] In some embodiments, the lysis buffer does not contain a detergent. In some embodiments, the lysis buffer does not contain a solubilizer.

[0142] The final volume of the lysis buffer may depend on the type of sample or amplification method. In some embodiments, the final volume of the lysis buffer is at least about 50 microliterWSGR Docket No. 52459-726.601(pl), at least about 100 pl, at least about 150 pl, at least about 200 pl, at least about 250 pl, at least about 300 pl, at least about 350 pl, at least about 400 pl, at least about 450 pl, at least about 500 pl, at least about 550 pl, at least about 600 pl, at least about 650 pl, at least about 700 pl, at least about 750 pl, at least about 800 pl, at least about 850 pl, at least about 900 pl, at least about 950 pl, at least about 1000 pl, at least about 1100 pl, at least about 1200 pl, at least about 1300 pl, at least about 1400 pl, at least about 1500 pl, at least about 2000 pl, at least about 2500 pl, at least about 3000 pl, at least about 4000 pl, at least about 5000 pl, at least about 7500 pl, or at least about 10,000 pl.

[0143] In some embodiments, the final volume of the lysis buffer is at most about 10,000 pl, at most about 7500 pl, at most about 5000 pl, at most about 4000 pl, at most about 3000 pl, at most about 2500 pl, at most about 2000 pl, at most about 1500 pl, at most about 1400 pl, at most about 1300 pl, at most about 1200 pl, at most about 1100 pl, at most about 1000 pl, at most about 950 pl, at most about 900 pl, at most about 850 pl, at most about 800 pl, at most about 750 pl, at most about 700 pl, at most about 650 pl, at most about 600 pl, at most about 550 pl, at most about 500 pl, at most about 450 pl, at most about 400 pl, at most about 350 pl, at most about 300 pl, at most about 250 pl, at most about 200 pl, at most about 150 pl, at most about 100 pl, or at most about 50 pl.

[0144] In some embodiments, the final volume of the lysis buffer is about 50 pl to about 2,000 pl. In some embodiments, the final volume of the lysis buffer is at most about 2,000 pl. In some embodiments, the final volume of the lysis buffer is about 50 pl to about 100 pl, about 50 pl to about 200 pl, about 50 pl to about 300 pl, about 50 pl to about 400 pl, about 50 pl to about 500 pl, about 50 pl to about 600 pl, about 50 pl to about 750 pl, about 50 pl to about 1,000 pl, about 50 pl to about 1,500 pl, about 50 pl to about 1,750 pl, about 50 pl to about 2,000 pl, about 100 pl to about 200 pl, about 100 pl to about 300 pl, about 100 pl to about 400 pl, about 100 pl to about 500 pl, about 100 pl to about 600 pl, about 100 pl to about 750 pl, about 100 pl to about 1,000 pl, about 100 pl to about 1,500 pl, about 100 pl to about 1,750 pl, about 100 pl to about 2,000 pl, about 200 pl to about 300 pl, about 200 pl to about 400 pl, about 200 pl to about 500 pl, about 200 pl to about 600 pl, about 200 pl to about 750 pl, about 200 pl to about 1,000 pl, about 200 pl to about 1,500 pl, about 200 pl to about 1,750 pl, about 200 pl to about 2,000 pl, about 300 pl to about 400 pl, about 300 pl to about 500 pl, about 300 pl to about 600 pl, about 300 pl to about 750 pl, about 300 pl to about 1,000 pl, about 300 pl to about 1,500 pl, about 300 pl to about 1,750 pl, about 300 pl to about 2,000 pl, about 400 pl to about 500 pl, about 400 pl to about 600 pl, about 400 pl to about 750 pl, about 400 pl to about 1,000 pl, about 400 pl toWSGR Docket No. 52459-726.601 about 1,500 pl, about 400 pl to about 1,750 pl, about 400 pl to about 2,000 pl, about 500 pl to about 600 pl, about 500 pl to about 750 pl, about 500 pl to about 1,000 pl, about 500 pl to about1,500 pl, about 500 pl to about 1,750 pl, about 500 pl to about 2,000 pl, about 600 pl to about 750 pl, about 600 pl to about 1,000 pl, about 600 pl to about 1,500 pl, about 600 pl to about 1,750 pl, about 600 pl to about 2,000 pl, about 750 pl to about 1,000 pl, about 750 pl to about1,500 pl, about 750 pl to about 1,750 pl, about 750 pl to about 2,000 pl, about 1,000 pl to about1,500 pl, about 1,000 pl to about 1,750 pl, about 1,000 pl to about 2,000 pl, about 1,500 pl to about 1,750 pl, about 1,500 pl to about 2,000 pl, or about 1,750 pl to about 2,000 pl.

[0145] In some embodiments, the final volume of the lysis buffer is about 2 ml to about 10 ml.

[0146] In some embodiments, the lysis buffer is lyophilized. In some embodiments, the lysis buffer is not lyophilized. In some embodiments, the final volume of the lysis buffer is about 2 ml to about 2.5 ml, about 2 ml to about 3 ml, about 2 ml to about 3.5 ml, about 2 ml to about 4 ml, about 2 ml to about 4.5 ml, about 2 ml to about 5 ml, about 2 ml to about 6 ml, about 2 ml to about 7 ml, about 2 ml to about 8 ml, about 2 ml to about 9 ml, about 2 ml to about 10 ml, about2.5 ml to about 3 ml, about 2.5 ml to about 3.5 ml, about 2.5 ml to about 4 ml, about 2.5 ml to about 4.5 ml, about 2.5 ml to about 5 ml, about 2.5 ml to about 6 ml, about 2.5 ml to about 7 ml, about 2.5 ml to about 8 ml, about 2.5 ml to about 9 ml, about 2.5 ml to about 10 ml, about 3 ml to about 3.5 ml, about 3 ml to about 4 ml, about 3 ml to about 4.5 ml, about 3 ml to about 5 ml, about 3 ml to about 6 ml, about 3 ml to about 7 ml, about 3 ml to about 8 ml, about 3 ml to about 9 ml, about 3 ml to about 10 ml, about 3.5 ml to about 4 ml, about 3.5 ml to about 4.5 ml, about3.5 ml to about 5 ml, about 3.5 ml to about 6 ml, about 3.5 ml to about 7 ml, about 3.5 ml to about 8 ml, about 3.5 ml to about 9 ml, about 3.5 ml to about 10 ml, about 4 ml to about 4.5 ml, about 4 ml to about 5 ml, about 4 ml to about 6 ml, about 4 ml to about 7 ml, about 4 ml to about 8 ml, about 4 ml to about 9 ml, about 4 ml to about 10 ml, about 4.5 ml to about 5 ml, about 4.5 ml to about 6 ml, about 4.5 ml to about 7 ml, about 4.5 ml to about 8 ml, about 4.5 ml to about 9 ml, about 4.5 ml to about 10 ml, about 5 ml to about 6 ml, about 5 ml to about 7 ml, about 5 ml to about 8 ml, about 5 ml to about 9 ml, about 5 ml to about 10 ml, about 6 ml to about 7 ml, about 6 ml to about 8 ml, about 6 ml to about 9 ml, about 6 ml to about 10 ml, about 7 ml to about 8 ml, about 7 ml to about 9 ml, about 7 ml to about 10 ml, about 8 ml to about 9 ml, about 8 ml to about 10 ml, or about 9 ml to about 10 ml.

[0147] In some embodiments, the solubilizer is a non-ionic surfactant. In some embodiments, the solubilizer comprises a polysorbate. The polysorbate may be polyoxyethylene (20) sorbitan monooleate (e.g., polysorbate 80), polyoxyethylene (20) sorbitan monolaurate (e.g., polysorbateWSGR Docket No. 52459-726.60120), polyoxyethylene (20) sorbitan monopalmitate (e.g., polysorbate 40), polyoxyethylene (20) sorbitan monostearate (e.g., polysorbate 60), or a functional variant thereof. In some embodiments, the solubilizer is a Tergitol™ surfactant, a Triton™ surfactant, or a Igepal® surfactant. In some embodiments, the solubilizer is an alkoxylate or a cocamide. In some embodiments, the solubilizer is decyl glucoside, alkyl polyglycoside, lauryl glucoside, sorbitan tristearate, or a niosome. In some embodiments, the recovery buffer contains one, two, three, four, or more solubilizers. The solubilizer may mix with the detergent of the present composition. In some embodiments, the solubilizer is capable of forming micelles comprising the detergent of the present application.

[0148] In some embodiments, the solubilizer is polysorbate 80. In some embodiments, the concentration (e.g., final concentration) of the solubilizer in the mixture in the presence of the sample is at least about 0.05% v / v, at least about 0.1% v / v, at least about 0.5% v / v, at least about 1% v / v, at least about 5% v / v, at least about 10% v / v, at least about 15% v / v, at least about 20% v / v, at least about 22.5% v / v, at least about 25% v / v, at least about 27.5% v / v, at least about 30% v / v, at least about 32.5% v / v, at least about 35% v / v, at least about 37.5% v / v, at least about 40% v / v, at least about 42.5% v / v, at least about 45% v / v, at least about 47.5% v / v, at least about 50% v / v, at least about 52.5% v / v, at least about 55% v / v, at least about 57.5% v / v, at least about 60% v / v, at least about 70% v / v, or at least about 75% v / v.

[0149] In some embodiments, the concentration (e.g., final concentration) of the solubilizer in the mixture in the presence of the sample is at most about 75% v / v, at most about 70% v / v, at most about 65% v / v, at most about 60% v / v, at most about 57.5% v / v, at most about 55% v / v, at most about 52.5% v / v, at most about 50% v / v, at most about 47.5% v / v, at most about 45% v / v, at most about 42.5% v / v, at most about 40% v / v, at most about 37.5% v / v, at most about 35% v / v, at most about 32.5% v / v, at most about 30% v / v, at most about 27.5% v / v, at most about 25% v / v, at most about 22.5% v / v, at most about 20% v / v, at most about 15% v / v, at most about 10% v / v, at most about 5% v / v, at most about 1% v / v, at most about 0.5% v / v, at most about 0.1% v / v, or at most about 0.05% v / v.

[0150] In some embodiments, the concentration (e.g., final concentration) of the solubilizer in the mixture in the presence of the sample is about 0.1% v / v to about 80% v / v. In some embodiments, the concentration (e.g., final concentration) of the solubilizer in the mixture in the presence of the sample is about 0.1% v / v to about 5% v / v, about 0.1% v / v to about 10% v / v, about 0.1% v / v to about 15% v / v, about 0.1% v / v to about 20% v / v, about 0.1% v / v to about 25% v / v, about 0.1% v / v to about 30% v / v, about 0.1% v / v to about 40% v / v, about 0.1% v / v toWSGR Docket No. 52459-726.601 about 50% v / v, about 0.1% v / v to about 60% v / v, about 0.1% v / v to about 70% v / v, about 0.1% v / v to about 80% v / v, about 5% v / v to about 10% v / v, about 5% v / v to about 15% v / v, about 5% v / v to about 20% v / v, about 5% v / v to about 25% v / v, about 5% v / v to about 30% v / v, about 5% v / v to about 40% v / v, about 5% v / v to about 50% v / v, about 5% v / v to about 60% v / v, about 5% v / v to about 70% v / v, about 5% v / v to about 80% v / v, about 10% v / v to about 15% v / v, about10% v / v to about 20% v / v, about 10% v / v to about 25% v / v, about 10% v / v to about 30% v / v, about 10% v / v to about 40% v / v, about 10% v / v to about 50% v / v, about 10% v / v to about 60% v / v, about 10% v / v to about 70% v / v, about 10% v / v to about 80% v / v, about 15% v / v to about 20% v / v, about 15% v / v to about 25% v / v, about 15% v / v to about 30% v / v, about 15% v / v to about 40% v / v, about 15% v / v to about 50% v / v, about 15% v / v to about 60% v / v, about 15% v / v to about 70% v / v, about 15% v / v to about 80% v / v, about 20% v / v to about 25% v / v, about 20% v / v to about 30% v / v, about 20% v / v to about 40% v / v, about 20% v / v to about 50% v / v, about 20% v / v to about 60% v / v, about 20% v / v to about 70% v / v, about 20% v / v to about 80% v / v, about 25% v / v to about 30% v / v, about 25% v / v to about 40% v / v, about 25% v / v to about 50% v / v, about 25% v / v to about 60% v / v, about 25% v / v to about 70% v / v, about 25% v / v to about 80% v / v, about 30% v / v to about 40% v / v, about 30% v / v to about 50% v / v, about 30% v / v to about 60% v / v, about 30% v / v to about 70% v / v, about 30% v / v to about 80% v / v, about 40% v / v to about 50% v / v, about 40% v / v to about 60% v / v, about 40% v / v to about 70% v / v, about 40% v / v to about 80% v / v, about 50% v / v to about 60% v / v, about 50% v / v to about 70% v / v, about 50% v / v to about 80% v / v, about 60% v / v to about 70% v / v, about 60% v / v to about 80% v / v, or about 70% v / v to about 80% v / v.

[0151] In some embodiments, the composition comprises a cyclodextrin. The cyclodextrin is configured to form a complex with the detergent of the present application. Without wishing to be bound by theory, the complex formed between the cyclodextrin and detergent assists in stabilizing the enzyme in the composition. The cyclodextrin increases the efficiency of forming the complex. As a complexing agent, the cyclodextrin can increase the aqueous solubility of poorly soluble drugs and increase bioavailability and stability in solution. In some embodiments, the cyclodextrin comprises (2-hydroxypropyl) P-cyclodextrin, (2-hydroxypropyl) y-cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P-cyclodextrin, 2,6-Di-O-methyl- P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3 A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, or any combination thereof. The cyclodextrin can comprise an anionic cyclodextrin. The anionic cyclodextrin may compriseWSGR Docket No. 52459-726.601 carboxymethyl-a-cyclodextrin, carboxymethyl-P-cyclodextrin, succinyl-a-cyclodextrin, succinyl-P-cyclodextrin, succinyl-y-cyclodextrin, (2-carboxyl)-a-cyclodextrin, (2-carboxyl)-P- cyclodextrin, a-cyclodextrin phosphate, P-cyclodextrin phosphate, y-cyclodextrin phosphate, sulfobutylated P-cyclodextrin, a-cyclodextrin sulfate, P-cyclodextrin sulfate, y-cyclodextrin sulfate, carboxymethyl-y-cyclodextrin, (2-carboxyl)-y-cyclodextrin, sulfobutylated-a- cyclodextrin, succinyl-(2-hydroxypropyl)-P cyclodextrin, succinyl-(2-hydroxypropyl)-y cyclodextrin, sulfobutylated-y cyclodextrin, methyl-P-cyclodextrin, or any combination thereof. In some embodiments, the cyclodextrin in the recovery buffer can comprise two or more different cyclodextrin species described herein. For example, the cyclodextrin in the recovery buffer can comprise (2-hydroxypropyl) p-cyclodextrin and (2-hydroxypropyl) y-cyclodextrin. For another example, the cyclodextrin in the recovery buffer can comprise (2-hydroxypropyl) a- cyclodextrin and methyl-P-cyclodextrin. In some cases, the cyclodextrin in the recovery buffer can comprise (2-hydroxypropyl) P-cyclodextrin and methyl-P-cyclodextrin. In some cases, altering the molar substitution ratio of a particular modified cyclodextrin species (e.g., (2- hydroxypropyl) P-cyclodextrin, methyl-P-cyclodextrin, etc.) may improve reaction performance such as shortening time to result values, Ct values, or Cq values.

[0152] In some embodiments, the recovery buffer does not comprise a component in the lysis buffer. For example, the recovery buffer may not comprise a detergent or a reducing agent. In some cases, the recovery buffer may not comprise one or more agent selected from the group consisting of an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2- carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), and tetrahydropyran (THP).

[0153] In some embodiments, the cyclodextrin is present at a final concentration in the presence of the sample effective for isolating the detergent within the composition of the present invention. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is at least about 0.05 mM, at least about 0.1 mM, at least about 0.5 mM, at least about 1.0 mM, at least about 5.0 mM, at least about 10.0 mM, at least about 15.0 mM, at least about 20.0 mM, at least about 25.0 mM, at least about 30.0 mM, at least about 35.0 mM, at least about 40.0 mM, at least about 50.0 mM, at least about 55.0 mM, at least about 60.0 mM, at least about 65.0 mM, at least about 70.0 mM, at least about 75.0 mM, atWSGR Docket No. 52459-726.601 least about 80.0 mM, at least about 85.0 mM, at least about 90.0 mM, at least about 95.0 mM, at least about 100.0 mM, at least about 125.0 mM, at least about 150.0 mM, at least about 175.0 mM, at least about 200.0 mM, at least about 250.0 mM, or at least about 300.0 mM.

[0154] In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is at most about 300.0 mM, at most about 250.0 mM, at most about 200.0 mM, at most about 175.0 mM, at most about 150.0 mM, at most about 125.0 mM, at most about 100.0 mM, at most about 95.0 mM, at most about 90.0 mM, at most about 85.0 mM, at most about 80.0 mM, at most about 75.0 mM, at most about 70.0 mM, at most about 65.0 mM, at most about 60.0 mM, at most about 55.0 mM, at most about 50.0 mM, at most about 45.0 mM, at most about 40.0 mM, at most about 35.0 mM, at most about 30.0 mM, at most about 25.0 mM, at most about 20.0 mM, at most about 15.0 mM, at most about 10.0 mM, at most about 5.0 mM, at most about 1.0 mM, at most about 0.5 mM, at most about 0.1 mM, or at most about 0.05 mM.

[0155] In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is about 10 mM to about 300 mM. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is at least about 10 mM. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is at most about 300 mM. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is about 10 mM to about 20 mM, about 10 mM to about 25 mM, about 10 mM to about 30 mM, about 10 mM to about 32.5 mM, about 10 mM to about 35 mM, about 10 mM to about 37.5 mM, about 10 mM to about 40 mM, about 10 mM to about 50 mM, about 10 mM to about 100 mM, about 10 mM to about 200 mM, about 10 mM to about 300 mM, about 20 mM to about 25 mM, about 20 mM to about 30 mM, about 20 mM to about 32.5 mM, about 20 mM to about 35 mM, about 20 mM to about 37.5 mM, about 20 mM to about 40 mM, about 20 mM to about 50 mM, about 20 mM to about 100 mM, about 20 mM to about 200 mM, about 20 mM to about 300 mM, about 25 mM to about 30 mM, about 25 mM to about 32.5 mM, about 25 mM to about 35 mM, about 25 mM to about 37.5 mM, about 25 mM to about 40 mM, about 25 mM to about 50 mM, about 25 mM to about 100 mM, about 25 mM to about 200 mM, about 25 mM to about 300 mM, about 30 mM to about 32.5 mM, about 30 mM to about 35 mM, about 30 mM to about 37.5 mM, about 30 mM to about 40 mM, about 30 mM to about 50 mM, about 30 mM to about 100 mM, about 30 mM to about 200 mM, about 30 mM to about 300 mM, about 32.5 mM to about 35 mM, about 32.5WSGR Docket No. 52459-726.601 mM to about 37.5 mM, about 32.5 mM to about 40 mM, about 32.5 mM to about 50 mM, about 32.5 mM to about 100 mM, about 32.5 mM to about 200 mM, about 32.5 mM to about 300 mM, about 35 mM to about 37.5 mM, about 35 mM to about 40 mM, about 35 mM to about 50 mM, about 35 mM to about 100 mM, about 35 mM to about 200 mM, about 35 mM to about 300 mM, about 37.5 mM to about 40 mM, about 37.5 mM to about 50 mM, about 37.5 mM to about 100 mM, about 37.5 mM to about 200 mM, about 37.5 mM to about 300 mM, about 40 mM to about 50 mM, about 40 mM to about 100 mM, about 40 mM to about 200 mM, about 40 mM to about 300 mM, about 50 mM to about 100 mM, about 50 mM to about 200 mM, about 50 mM to about 300 mM, about 100 mM to about 200 mM, about 100 mM to about 300 mM, or about 200 mM to about 300 mM.

[0156] In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is about 0.1 mM to about 100 mM. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is at most about 100 mM. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is about 0.1 mM to about 1 mM, about 0.1 mM to about 5 mM, about 0.1 mM to about 10 mM, about 0.1 mM to about 20 mM, about 0.1 mM to about 30 mM, about 0.1 mM to about 35 mM, about 0.1 mM to about 40 mM, about 0.1 mM to about 50 mM, about 0.1 mM to about 60 mM, about 0.1 mM to about 75 mM, about 0.1 mM to about 100 mM, about 1 mM to about 5 mM, about 1 mM to about 10 mM, about 1 mM to about 20 mM, about 1 mM to about 30 mM, about 1 mM to about 35 mM, about 1 mM to about 40 mM, about 1 mM to about 50 mM, about 1 mM to about 60 mM, about 1 mM to about 75 mM, about 1 mM to about 100 mM, about 5 mM to about 10 mM, about 5 mM to about 20 mM, about 5 mM to about 30 mM, about 5 mM to about 35 mM, about 5 mM to about 40 mM, about 5 mM to about 50 mM, about 5 mM to about 60 mM, about 5 mM to about 75 mM, about 5 mM to about 100 mM, about 10 mM to about 20 mM, about 10 mM to about 30 mM, about 10 mM to about 35 mM, about 10 mM to about 40 mM, about 10 mM to about 50 mM, about 10 mM to about 60 mM, about 10 mM to about 75 mM, about 10 mM to about 100 mM, about 20 mM to about 30 mM, about 20 mM to about 35 mM, about 20 mM to about 40 mM, about 20 mM to about 50 mM, about 20 mM to about 60 mM, about 20 mM to about 75 mM, about 20 mM to about 100 mM, about 30 mM to about 35 mM, about 30 mM to about 40 mM, about 30 mM to about 50 mM, about 30 mM to about 60 mM, about 30 mM to about 75 mM, about 30 mM to about 100 mM, about 35 mM to about 40 mM, about 35 mM to about 50 mM, about 35 mM to about 60 mM, about 35 mM to about 75WSGR Docket No. 52459-726.601 mM, about 35 mM to about 100 mM, about 40 mM to about 50 mM, about 40 mM to about 60 mM, about 40 mM to about 75 mM, about 40 mM to about 100 mM, about 50 mM to about 60 mM, about 50 mM to about 75 mM, about 50 mM to about 100 mM, about 60 mM to about 75 mM, about 60 mM to about 100 mM, or about 75 mM to about 100 mM.

[0157] In some embodiments, the recovery buffer can comprise a cyclodextrin with a final concentration of about 6 mM to 11 mM in the presence of the sample and polysorbate 80 with a final concentration of about 0.1% v / v to 3.0% v / v in the presence of the sample. For example, the recovery buffer can comprise 2 mM cyclodextrin and 1.5% v / v polysorbate 80 final concentration for each component in the presence of the sample.

[0158] In some embodiments, the cyclodextrin has a higher binding affinity toward the detergent than a binding affinity of the solubilizer towards the detergent. In some embodiments, the binding affinity of the cyclodextrin to the detergent can be an association constant. In some embodiments, the binding affinity of the cyclodextrin to the detergent has an association constant (Ka) of at least about 2.5xl03M’1, at least about 3xl03M’1, at least about 3.5xl03M’1, at least about 4xl03M’1, at least about 5xl03M’1, at least about IxlO4M’1, at least about 2xl04M’1, at least about 3xl04M’1, at least about 4xl04M’1, at least about 5xl04M’1, at least about IxlO5M’1, at least about 5xl05M’1, or at least about IxlO6M'1to the detergent. In some embodiments, the binding affinity of the cyclodextrin to the detergent has an association constant (Ka) of at most about IxlO6M’1, at most about 5xl05M’1, at most about IxlO5M’1, at most about 5xl04M’1, at most about 4xl04M’1, at most about 3xl04M’1, at most about 2xl04M’1, at most about IxlO4M’1, at most about 5xl03M’1, at most about 4xl03M’1, at most about 3xl03M’1, or at most about 2.5xl03M'1.

[0159] In some embodiments, the solubilizer and the cyclodextrin are configured to shorten a cycle threshold value or time to result value in a nucleic acid amplification compared to a cycle threshold value or a time to result value in an nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually. The term “cycle threshold” refers to the number of cycles used to amplify a target nucleic acid molecule to a detectable level (e.g., a signal exceeding a background threshold level). A lower cycle threshold value can indicate a greater amount of target nucleic acid in a sample. In some cases, when isothermal amplifications are used, time to result value can also be used and it refers to the time used to amplify a target nucleic acid molecule to a detectable level. In some embodiments, the solubilizer and / or cyclodextrin described herein are configured to shorten a cycle threshold value to at most about 60, at most about 50, at most about 40, at most about 30, at most aboutWSGR Docket No. 52459-726.60125, at most about 20, at most about 19, at most about 18, at most about 17, at most about 16, at most about 15, at most about 14, at most about 13, at most about 12, at most about 11, at most about 10, at most about 9, at most about 8, at most about 7, at most about 6, at most about 5, at most about 4, at most about 3, at most about 2, or at most about 1. In some embodiments, the solubilizer and / or cyclodextrin described herein are configured to shorten a time to result value to at most about 15 minutes, at most about 14 minutes, at most about 13 minutes, at most about 12 minutes, at most about 11 minutes, at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6 minutes, at most about 5 minutes or less.

[0160] In some embodiments, the solubilizer and / or the cyclodextrin are configured to decrease a coefficient of variation in a nucleic acid amplification compared to a coefficient of variation in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually. The term “coefficient of variation” refers to a measure of precision of an amplification method. In some embodiments, the solubilizer and / or the cyclodextrin are configured to decrease a coefficient of variation value to at most about 15%, at most about 14%, at most about 13%, at most about 12%, at most about 11%, at most about 10%, at most about 9%, at most about 8%, at most about 7%, at most about 6%, at most about 5%, at most about 4.5%, at most about 4%, at most about 3.5%, at most about 3%, at most about 2.5%, at most about 2%, at most about 1.5%, or at most about 1%.

[0161] In some embodiments, the solubilizer and / or the cyclodextrin are configured to lower a limit of detection of a nucleic acid amplification compared to a limit of detection in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually. The “limit of detection” refers to the lowest quantity of a component in a sample that be reliably detected in an amplification method. In some embodiments, the solubilizer and / or the cyclodextrin are configured to lower a limit of detection to about 1 target molecule, about 1.5 target molecules, about 2 target molecules, about 2.5 target molecules, about 3 target molecules, about 3.5 target molecules, about 4 target molecules, about 4.5 target molecules, about 5 target molecules, about 6 target molecules, about 7 target molecules, about 8 target molecules, about 9 target molecules, or about 10 target molecules.

[0162] In some embodiments, the solubilizer and the cyclodextrin are part of a recovery buffer. In some embodiments, the recovery buffer comprises a salt. In some embodiments, the recovery buffer does not comprise a salt. In some embodiments, the salt comprises a sodium salt. In some embodiments, the recovery buffer comprises a pH buffer. In some embodiments, the recoveryWSGR Docket No. 52459-726.601 buffer does not comprise a pH buffer. In some embodiments, the pH of the recovery buffer is at least about 3, at least about 4, at least about 4.5, at least about 5, at least about 5.5, at least about 6, at least about 6.5, at least about 7, at least about 7.5, at least about 8, at least about 9, at least about 10, at least about 11, or at least about 12. In some embodiments, the pH of the recovery buffer is at most about 12, at most about 11, at most about 10, at most about 9, at most about 8, at most about 7.5, at most about 7, at most about 6.5, at most about 6, at most about 5.5, at most about 5, at most about 4.5, at most about 4, or at most about 3.

[0163] In some embodiments, the recovery buffer is lyophilized. The recovery buffer can be liquid. The recovery buffer can be lyophilized together with a reaction buffer / reaction mixture for nucleic acid amplifications.

[0164] In some embodiments, the lysis buffer and the recovery buffer are in the same mixture. In some embodiments, the mixing of the lysis buffer and the recovery buffer is performed by hand. In some embodiments, the mixing of the lysis buffer and the recovery buffer is performed by a vortex. In some embodiments, the mixing of the lysis buffer and the recovery buffer is performed by an automated instrument, a consumable, or a microfluidic system. In some embodiments, the mixing of the lysis buffer and the recovery buffer is performed until the lysis buffer and the recovery buffer are mixed to homogeneity.

[0165] In some aspects, the present disclosure provides for a composition for sample processing comprising a buffer comprising: (i) a detergent), (ii) a solubilizer, and (iii) a cyclodextrin. In some embodiments, the buffer stabilizes an enzyme during a nucleic acid amplification. In some embodiments, the buffer is configured to inactivate a degrading enzyme. In some embodiments, the enzyme is a ribonuclease.

[0166] In some embodiments, the composition of the present disclosure further comprises an agent capable of reducing a disulfide bond. In some embodiments, the agent capable of reducing said disulfide bond comprises dithiothreitol (DTT), hydroxylamine, hydroxylamine-HCl, 2- mercaptoethanol (BME), or TCEP. In some embodiments, the agent capable of reducing said disulfide bond comprises a compound in a monothiol class, a dithiol class, or a phosphine class.

[0167] In some embodiments, the composition further comprises a sample, (e.g., a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, and / or a lymph sample), raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or moreWSGR Docket No. 52459-726.601 whole organisms, one or more homogenized organisms, wastewater, or any combination thereof for nucleic acid amplification. In some embodiments, the swab sample comprises a vaginal swab, an oral swab, and / or a rectal swab. In some embodiments, the sample is a solid sample. In some embodiments, the sample is a liquid sample. In some embodiments, the sample is obtained from a subject. In some embodiments, the subject has a disease, a condition, or an infection. In some embodiments, the sample comprises a biological sample. In some embodiments, the sample comprises a purified sample. In some embodiments, the biological sample comprises a target nucleic acid molecule subject to sample processing.

[0168] In some embodiments, the composition further comprises a reaction mixture for nucleic acid amplification. In some embodiments, the reaction mixture is lyophilized. In some embodiments, the reaction mixture is not lyophilized. In some embodiments, the reaction mixture comprises (i) a thermostable enzyme, (ii) deoxynucleoside triphosphates (dNTPs), (iii) a primer, and / or (iv) a probe. In some embodiments, the thermostable enzyme comprises a Bacillus stearothermophilus polymerase, a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, a Thermus aquaticus (e.g., Taq-polA), a Thermotoga maritima (e.g., Tma-polA), a Pfu-polB, a Pab-polB, an OmniTaq 2 LA DNA polymerase, or any mutants thereof. A large fragment of a Bacillus stearothermophilus polymerase is the portion of the Bacillus stearothermophilus DNA polymerase that contains the 5' — > 3' polymerase activity, but lacks the 5' -^ ' exonuclease domain. In some embodiments, the composition is configured to stabilize enzymatic activity of the thermostable enzyme for use during a nucleic acid amplification.

[0169] In some embodiments, the dNTPs of the reaction mixture comprise dATP, dCTP, dGTP, dTTP, and / or dUTP. In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is at least about 25 micromolar (pM), at least about 50 pM, at least about 75 pM, at least about 100 pM, at least about 150 pM, at least about 200 pM, at least about 250 pM, at least about 300 pM, at least about 350 pM, at least about 400 pM, at least about 450 pM, at least about 500 pM, at least about 750 pM, at least about 1000 pM, at least about 1500 pM, at least about 2000 pM, at least about 2500 pM, at least about 3000 pM, at least about 3500 pM, at least about 4000 pM, at least about 4500 pM, at least about 5000 pM, at least about 6000 pM, at least about 7000 pM, at least about 8000 pM, at least about 9000 pM, or at least about 10000 pM.WSGR Docket No. 52459-726.601

[0170] In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is at most about 10000 pM, at most about 9000 pM, at most about 8000 pM, at most about 7000 pM, at most about 6000 pM, at most about 5000 pM, at most about 4500 pM, at most about 4000 pM, at most about 3500 pM, at most about 3000 pM, at most about 2500 pM, at most about 2000 pM, at most about 1500 pM, at most about 1000 pM, at most about 750 pM, at most about 500 pM, at most about 450 pM, at most about 400 pM, at most about 350 pM, at most about 300 pM, at most about 250 pM, at most about 200 pM, at most about 150 pM, at most about 100 pM, at most about 75 pM, at most about 50 pM, or at most about 25 pM.

[0171] In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is about 50 pM to about 7,500 pM. In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is about 50 pM to about 100 pM, about 50 pM to about 250 pM, about 50 pM to about 500 pM, about 50 pM to about 750 pM, about 50 pM to about 1,000 pM, about 50 pM to about 1,250 pM, about 50 pM to about 1,500 pM, about 50 pM to about 2,000 pM, about 50 pM to about 4,000 pM, about 50 pM to about 5,000 pM, about 50 pM to about 7,500 pM, about 100 pM to about 250 pM, about 100 pM to about 500 pM, about 100 pM to about 750 pM, about 100 pM to about 1,000 pM, about 100 pM to about 1,250 pM, about 100 pM to about 1,500 pM, about 100 pM to about 2,000 pM, about 100 pM to about 4,000 pM, about 100 pM to about 5,000 pM, about 100 pM to about 7,500 pM, about 250 pM to about 500 pM, about 250 pM to about 750 pM, about 250 pM to about 1,000 pM, about 250 pM to about 1,250 pM, about 250 pM to about 1,500 pM, about 250 pM to about 2,000 pM, about 250 pM to about 4,000 pM, about 250 pM to about 5,000 pM, about 250 pM to about 7,500 pM, about 500 pM to about 750 pM, about 500 pM to about 1,000 pM, about 500 pM to about 1,250 pM, about 500 pM to about 1,500 pM, about 500 pM to about 2,000 pM, about 500 pM to about 4,000 pM, about 500 pM to about 5,000 pM, about 500 pM to about 7,500 pM, about 750 pM to about 1,000 pM, about 750 pM to about 1,250 pM, about 750 pM to about 1,500 pM, about 750 pM to about 2,000 pM, about 750 pM to about 4,000 pM, about 750 pM to about 5,000 pM, about 750 pM to about 7,500 pM, about 1,000 pM to about 1,250 pM, about 1,000 pM to about 1,500 pM, about 1,000 pM to about 2,000 pM, about 1,000 pM to about 4,000 pM, about 1,000 pM to about 5,000 pM, about 1,000 pM to about 7,500 pM, about 1,250 pM to about 1,500 pM, about 1,250 pM to about 2,000 pM, about 1,250 pM to about 4,000 pM, about 1,250 pM to about 5,000 pM, about 1,250 pM to about 7,500 pM, about 1,500 pM to about 2,000 pM, about 1,500 pM to about 4,000 pM, about 1,500 pM to about 5,000 pM, about 1,500 pM to about 7,500 pM, about 2,000 pM to about 4,000 pM,WSGR Docket No. 52459-726.601 about 2,000 pM to about 5,000 pM, about 2,000 pM to about 7,500 pM, about 4,000 pM to about 5,000 pM, about 4,000 pM to about 7,500 pM, or about 5,000 pM to about 7,500 pM.

[0172] In some embodiments, the primer or probe can be a stretch of nucleotides that hybridizes with a target nucleic acid sequence. In some embodiments, the primer is at least about 3 nucleotides, at least about 5 nucleotides, at least about 10 nucleotides, at least about 15 nucleotides, at least about 20 nucleotides, at least about 25 nucleotides, at least about 30 nucleotides, at least about 35 nucleotides, at least about 40 nucleotides, at least about 45 nucleotides, at least about 50 nucleotides, at least about 60 nucleotides, at least about 70 nucleotides, at least about 80 nucleotides, at least about 90 nucleotides, at least about 100 nucleotides, at least about 150 nucleotides, or at least about 200 nucleotides in length. In some embodiments, the primer is at most about 200 nucleotides, at most about 150 nucleotides, at most about 100 nucleotides, at most about 90 nucleotides, at most about 80 nucleotides, at most about 70 nucleotides, at most about 60 nucleotides, at most about 50 nucleotides, at most about 45 nucleotides, at most about 40 nucleotides, at most about 35 nucleotides, at most about 30 nucleotides, at most about 25 nucleotides, at most about 20 nucleotides, at most about 15 nucleotides, at most about 10 nucleotides, at most about 5 nucleotides, or at most about 3 nucleotides in length.

[0173] In some embodiments, the primer is about 3 nucleotides to about 100 nucleotides in length. In some embodiments, the primer is at most about 100 nucleotides. In some embodiments, the primer is about 3 nucleotides to about 5 nucleotides, about 3 nucleotides to about 10 nucleotides, about 3 nucleotides to about 20 nucleotides, about 3 nucleotides to about 30 nucleotides, about 3 nucleotides to about 40 nucleotides, about 3 nucleotides to about 50 nucleotides, about 3 nucleotides to about 60 nucleotides, about 3 nucleotides to about 70 nucleotides, about 3 nucleotides to about 80 nucleotides, about 3 nucleotides to about 90 nucleotides, about 3 nucleotides to about 100 nucleotides, about 5 nucleotides to about 10 nucleotides, about 5 nucleotides to about 20 nucleotides, about 5 nucleotides to about 30 nucleotides, about 5 nucleotides to about 40 nucleotides, about 5 nucleotides to about 50 nucleotides, about 5 nucleotides to about 60 nucleotides, about 5 nucleotides to about 70 nucleotides, about 5 nucleotides to about 80 nucleotides, about 5 nucleotides to about 90 nucleotides, about 5 nucleotides to about 100 nucleotides, about 10 nucleotides to about 20 nucleotides, about 10 nucleotides to about 30 nucleotides, about 10 nucleotides to about 40 nucleotides, about 10 nucleotides to about 50 nucleotides, about 10 nucleotides to about 60 nucleotides, about 10 nucleotides to about 70 nucleotides, about 10 nucleotides to about 80WSGR Docket No. 52459-726.601 nucleotides, about 10 nucleotides to about 90 nucleotides, about 10 nucleotides to about 100 nucleotides, about 20 nucleotides to about 30 nucleotides, about 20 nucleotides to about 40 nucleotides, about 20 nucleotides to about 50 nucleotides, about 20 nucleotides to about 60 nucleotides, about 20 nucleotides to about 70 nucleotides, about 20 nucleotides to about 80 nucleotides, about 20 nucleotides to about 90 nucleotides, about 20 nucleotides to about 100 nucleotides, about 30 nucleotides to about 40 nucleotides, about 30 nucleotides to about 50 nucleotides, about 30 nucleotides to about 60 nucleotides, about 30 nucleotides to about 70 nucleotides, about 30 nucleotides to about 80 nucleotides, about 30 nucleotides to about 90 nucleotides, about 30 nucleotides to about 100 nucleotides, about 40 nucleotides to about 50 nucleotides, about 40 nucleotides to about 60 nucleotides, about 40 nucleotides to about 70 nucleotides, about 40 nucleotides to about 80 nucleotides, about 40 nucleotides to about 90 nucleotides, about 40 nucleotides to about 100 nucleotides, about 50 nucleotides to about 60 nucleotides, about 50 nucleotides to about 70 nucleotides, about 50 nucleotides to about 80 nucleotides, about 50 nucleotides to about 90 nucleotides, about 50 nucleotides to about 100 nucleotides, about 60 nucleotides to about 70 nucleotides, about 60 nucleotides to about 80 nucleotides, about 60 nucleotides to about 90 nucleotides, about 60 nucleotides to about 100 nucleotides, about 70 nucleotides to about 80 nucleotides, about 70 nucleotides to about 90 nucleotides, about 70 nucleotides to about 100 nucleotides, about 80 nucleotides to about 90 nucleotides, about 80 nucleotides to about 100 nucleotides, or about 90 nucleotides to about 100 nucleotides in length.

[0174] In some embodiments, the reaction mixture includes probes to visualize amplified nucleic acid products. In some embodiments, the probes comprise strand displacement probes, intercalating fluorophores, pH-sensitive dyes, and / or detecting pyrophosphate products.

[0175] The reaction mixture described herein can comprise an excipient. The excipient can comprise a saccharide (e.g., a monosaccharide, a disaccharide, a polysaccharide, or any combination thereof). The excipient can comprise a surfactant (e.g., nonoxynol-9). In some embodiments, the excipient comprises a polymer comprising a cross-linking sucrose with epichlorohydrin. The polymer may be polysucrose 400. In some embodiments, the excipient comprises Tris, potassium phosphate, sodium chloride, ethylenediaminetetraacetic acid (EDTA), potassium chloride, nonoxynol-9, trehalose, dextran, polysucrose 400, a cyclodextrin, or any combination thereof. The excipient may comprise dithiothreitol (DTT).

[0176] The cyclodextrin of the excipient can be hydroxypropyl P-cyclodextrin, hydroxypropyl y-cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a-WSGR Docket No. 52459-726.601 cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P- cyclodextrin, 2,6-Di-O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A- deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, an anionic cyclodextrin, or any combination thereof.

[0177] In some embodiments, the excipient may comprise a final concentration of Tris, sodium chloride and / or potassium chloride, EDTA, nonoxynol-9, one or more saccharides (e.g., dextran and / or trehalose), polysucrose 400, and / or cyclodextrin.

[0178] In some embodiments, the concentration (e.g., final concentration) of a Tris in the excipient is at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, at least about 100 mM, at least about 250 mM, at least about 500 mM, at least about 750 mM, at least about 1000 mM, at least about 1500 mM, at least about 2000 mM, or greater than about 2000 mM.

[0179] In some embodiments, the concentration (e.g., final concentration) of a Tris in the excipient is at most about 2000 mM, at most about 1500 mM, at most about 1000 mM, at most about 750 mM, at most about 500 mM, at most about 250 mM, at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, or at most about 1 mM.

[0180] In some embodiments, the concentration (e.g., final concentration) of a Tris in the excipient can be between about 0.0001 M to about 5 M. In some embodiments, the concentration (e.g., final concentration) of a Tris in the excipient can be between at most about 5 M. In some embodiments, the concentration (e.g., final concentration) of a Tris in the excipient can be between about 0.0001 M to about 0.0005 M, about 0.0001 M to about 0.001 M, aboutWSGR Docket No. 52459-726.6010.0001 M to about 0.005 M, about 0.0001 M to about 0.0075 M, about 0.0001 M to about 0.01 M, about 0.0001 M to about 0.025 M, about 0.0001 M to about 0.05 M, about 0.0001 M to about 0.1 M, about 0.0001 M to about 0.5 M, about 0.0001 M to about 1 M, about 0.0001 M to about 5 M, about 0.0005 M to about 0.001 M, about 0.0005 M to about 0.005 M, about 0.0005 M to about 0.0075 M, about 0.0005 M to about 0.01 M, about 0.0005 M to about 0.025 M, about 0.0005 M to about 0.05 M, about 0.0005 M to about 0.1 M, about 0.0005 M to about 0.5 M, about 0.0005 M to about 1 M, about 0.0005 M to about 5 M, about 0.001 M to about 0.005 M, about 0.001 M to about 0.0075 M, about 0.001 M to about 0.01 M, about 0.001 M to about 0.025 M, about 0.001 M to about 0.05 M, about 0.001 M to about 0.1 M, about 0.001 M to about 0.5 M, about 0.001 M to about 1 M, about 0.001 M to about 5 M, about 0.005 M to about 0.0075 M, about 0.005 M to about 0.01 M, about 0.005 M to about 0.025 M, about 0.005 M to about 0.05 M, about 0.005 M to about 0.1 M, about 0.005 M to about 0.5 M, about 0.005 M to about 1 M, about 0.005 M to about 5 M, about 0.0075 M to about 0.01 M, about 0.0075 M to about 0.025 M, about 0.0075 M to about 0.05 M, about 0.0075 M to about 0.1 M, about 0.0075 M to about 0.5 M, about 0.0075 M to about 1 M, about 0.0075 M to about 5 M, about 0.01 M to about 0.025 M, about 0.01 M to about 0.05 M, about 0.01 M to about 0.1 M, about 0.01 M to about 0.5 M, about 0.01 M to about 1 M, about 0.01 M to about 5 M, about 0.025 M to about 0.05 M, about 0.025 M to about 0.1 M, about 0.025 M to about 0.5 M, about 0.025 M to about 1 M, about 0.025 M to about 5 M, about 0.05 M to about 0.1 M, about 0.05 M to about 0.5 M, about 0.05 M to about 1 M, about 0.05 M to about 5 M, about 0.1 M to about 0.5 M, about 0.1 M to about 1 M, about 0.1 M to about 5 M, about 0.5 M to about 1 M, about 0.5 M to about 5 M, or about 1 M to about 5 M.

[0181] In some embodiments, the concentration (e.g., final concentration) of sodium chloride and / or potassium chloride in the excipient is at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, at least about 100 mM, at least about 250 mM, at least about 500 mM, at least about 750 mM, at least about 1000 mM, at least about 1500 mM, at least about 2000 mM, or greater than about 2000 mM.WSGR Docket No. 52459-726.601

[0182] In some embodiments, the concentration (e.g., final concentration) of sodium chloride and / or potassium chloride in the excipient is at most about 2000 mM, at most about 1500 mM, at most about 1000 mM, at most about 750 mM, at most about 500 mM, at most about 250 mM, at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, or at most about 1 mM.

[0183] In some embodiments, the concentration (e.g., final concentration) of sodium chloride and / or potassium chloride in the excipient can be between about 0.0001 M to about 5 M. In some embodiments, the concentration (e.g., final concentration) of sodium chloride and / or potassium chloride in the excipient can be between at most about 5 M. In some embodiments, the concentration (e.g., final concentration) of sodium chloride and / or potassium chloride in the excipient can be between about 0.0001 M to about 0.0005 M, about 0.0001 M to about 0.001 M, about 0.0001 M to about 0.005 M, about 0.0001 M to about 0.0075 M, about 0.0001 M to about 0.01 M, about 0.0001 M to about 0.025 M, about 0.0001 M to about 0.05 M, about 0.0001 M to about 0.1 M, about 0.0001 M to about 0.5 M, about 0.0001 M to about 1 M, about 0.0001 M to about 5 M, about 0.0005 M to about 0.001 M, about 0.0005 M to about 0.005 M, about 0.0005 M to about 0.0075 M, about 0.0005 M to about 0.01 M, about 0.0005 M to about 0.025 M, about 0.0005 M to about 0.05 M, about 0.0005 M to about 0.1 M, about 0.0005 M to about 0.5 M, about 0.0005 M to about 1 M, about 0.0005 M to about 5 M, about 0.001 M to about 0.005 M, about 0.001 M to about 0.0075 M, about 0.001 M to about 0.01 M, about 0.001 M to about 0.025 M, about 0.001 M to about 0.05 M, about 0.001 M to about 0.1 M, about 0.001 M to about 0.5 M, about 0.001 M to about 1 M, about 0.001 M to about 5 M, about 0.005 M to about 0.0075 M, about 0.005 M to about 0.01 M, about 0.005 M to about 0.025 M, about 0.005 M to about 0.05 M, about 0.005 M to about 0.1 M, about 0.005 M to about 0.5 M, about 0.005 M to about 1 M, about 0.005 M to about 5 M, about 0.0075 M to about 0.01 M, about 0.0075 M to about 0.025 M, about 0.0075 M to about 0.05 M, about 0.0075 M to about 0.1 M, about 0.0075 M to about 0.5 M, about 0.0075 M to about 1 M, about 0.0075 M to about 5 M, about 0.01 M to about 0.025 M, about 0.01 M to about 0.05 M, about 0.01 M to about 0.1 M, about 0.01 M to about 0.5WSGR Docket No. 52459-726.601M, about 0.01 M to about 1 M, about 0.01 M to about 5 M, about 0.025 M to about 0.05 M, about 0.025 M to about 0.1 M, about 0.025 M to about 0.5 M, about 0.025 M to about 1 M, about 0.025 M to about 5 M, about 0.05 M to about 0.1 M, about 0.05 M to about 0.5 M, about 0.05 M to about 1 M, about 0.05 M to about 5 M, about 0.1 M to about 0.5 M, about 0.1 M to about 1 M, about 0.1 M to about 5 M, about 0.5 M to about 1 M, about 0.5 M to about 5 M, or about 1 M to about 5 M.

[0184] In some embodiments, the concentration (e.g., final concentration) of dithiothreitol (DTT) in the excipient is at least about 0.01 mM, at least about 0.05 mM, at least about 0.1 mM, at least about 0.5 mM, at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, at least about 100 mM, at least about 250 mM, at least about 500 mM, at least about 750 mM, at least about 1000 mM, at least about 1500 mM, at least about 2000 mM, or greater than about 2000 mM.

[0185] In some embodiments, the concentration (e.g., final concentration) of dithiothreitol (DTT) in the excipient is at most about 2000 mM, at most about 1500 mM, at most about 1000 mM, at most about 750 mM, at most about 500 mM, at most about 250 mM, at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, at most about 1 mM, at most about 0.5 mM, at most about 0.1 mM, at most about 0.05 mM, at most about 0.01 mM, or less than about 0.01 mM.

[0186] In some embodiments, the concentration (e.g., final concentration) of dithiothreitol (DTT) in the excipient can be between about 0.0001 M to about 5 M. In some embodiments, the concentration (e.g., final concentration) of dithiothreitol (DTT) in the excipient can be betweenWSGR Docket No. 52459-726.601 at most about 5 M. In some embodiments, the concentration (e.g., final concentration) of dithiothreitol (DTT) in the excipient can be between about 0.0001 M to about 0.0005 M, about 0.0001 M to about 0.001 M, about 0.0001 M to about 0.005 M, about 0.0001 M to about 0.0075 M, about 0.0001 M to about 0.01 M, about 0.0001 M to about 0.025 M, about 0.0001 M to about 0.05 M, about 0.0001 M to about 0.1 M, about 0.0001 M to about 0.5 M, about 0.0001 M to about 1 M, about 0.0001 M to about 5 M, about 0.0005 M to about 0.001 M, about 0.0005 M to about 0.005 M, about 0.0005 M to about 0.0075 M, about 0.0005 M to about 0.01 M, about 0.0005 M to about 0.025 M, about 0.0005 M to about 0.05 M, about 0.0005 M to about 0.1 M, about 0.0005 M to about 0.5 M, about 0.0005 M to about 1 M, about 0.0005 M to about 5 M, about 0.001 M to about 0.005 M, about 0.001 M to about 0.0075 M, about 0.001 M to about 0.01 M, about 0.001 M to about 0.025 M, about 0.001 M to about 0.05 M, about 0.001 M to about 0.1 M, about 0.001 M to about 0.5 M, about 0.001 M to about 1 M, about 0.001 M to about 5 M, about 0.005 M to about 0.0075 M, about 0.005 M to about 0.01 M, about 0.005 M to about 0.025 M, about 0.005 M to about 0.05 M, about 0.005 M to about 0.1 M, about 0.005 M to about 0.5 M, about 0.005 M to about 1 M, about 0.005 M to about 5 M, about 0.0075 M to about 0.01 M, about 0.0075 M to about 0.025 M, about 0.0075 M to about 0.05 M, about 0.0075 M to about 0.1 M, about 0.0075 M to about 0.5 M, about 0.0075 M to about 1 M, about 0.0075 M to about 5 M, about 0.01 M to about 0.025 M, about 0.01 M to about 0.05 M, about 0.01 M to about 0.1 M, about 0.01 M to about 0.5 M, about 0.01 M to about 1 M, about 0.01 M to about 5 M, about 0.025 M to about 0.05 M, about 0.025 M to about 0.1 M, about 0.025 M to about 0.5 M, about 0.025 M to about 1 M, about 0.025 M to about 5 M, about 0.05 M to about 0.1 M, about 0.05 M to about 0.5 M, about 0.05 M to about 1 M, about 0.05 M to about 5 M, about 0.1 M to about 0.5 M, about 0.1 M to about 1 M, about 0.1 M to about 5 M, about 0.5 M to about 1 M, about 0.5 M to about 5 M, or about 1 M to about 5 M.

[0187] In some embodiments, the concentration (e.g., final concentration) of EDTA in the excipient is at least about 0.01 mM, at least about 0.05 mM, at least about 0.1 mM, at least about 0.5 mM, at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, atWSGR Docket No. 52459-726.601 least about 100 mM, at least about 250 mM, at least about 500 mM, at least about 750 mM, at least about 1000 mM, at least about 1500 mM, at least about 2000 mM, or greater than about 2000 mM.

[0188] In some embodiments, the concentration (e.g., final concentration) of EDTA in the excipient is at most about 2000 mM, at most about 1500 mM, at most about 1000 mM, at most about 750 mM, at most about 500 mM, at most about 250 mM, at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, at most about 1 mM, at most about 0.5 mM, at most about 0.1 mM, at most about 0.05 mM, at most about 0.01 mM, or less than about 0.01 mM.

[0189] In some embodiments, the concentration (e.g., final concentration) of EDTA in the excipient can be between about 0.01 mM to about 5 mM. In some embodiments, the concentration (e.g., final concentration) of EDTA in the excipient can be between at most about 5 mM. In some embodiments, the concentration (e.g., final concentration) of EDTA in the excipient can be between about 0.01 mM to about 0.05 mM, about 0.01 mM to about 0.1 mM, about 0.01 mM to about 0.5 mM, about 0.01 mM to about 0.75 mM, about 0.01 mM to about 1 mM, about 0.01 mM to about 1.25 mM, about 0.01 mM to about 1.5 mM, about 0.01 mM to about 1.75 mM, about 0.01 mM to about 2 mM, about 0.01 mM to about 3 mM, about 0.01 mM to about 5 mM, about 0.05 mM to about 0.1 mM, about 0.05 mM to about 0.5 mM, about 0.05 mM to about 0.75 mM, about 0.05 mM to about 1 mM, about 0.05 mM to about 1.25 mM, about 0.05 mM to about 1.5 mM, about 0.05 mM to about 1.75 mM, about 0.05 mM to about 2 mM, about 0.05 mM to about 3 mM, about 0.05 mM to about 5 mM, about 0.1 mM to about 0.5 mM, about 0.1 mM to about 0.75 mM, about 0.1 mM to about 1 mM, about 0.1 mM to about 1.25 mM, about 0.1 mM to about 1.5 mM, about 0.1 mM to about 1.75 mM, about 0.1 mM to about 2 mM, about 0.1 mM to about 3 mM, about 0.1 mM to about 5 mM, about 0.5 mM to about 0.75 mM, about 0.5 mM to about 1 mM, about 0.5 mM to about 1.25 mM, about 0.5 mM to about 1.5 mM, about 0.5 mM to about 1.75 mM, about 0.5 mM to about 2 mM, about 0.5 mM to about 3 mM, about 0.5 mM to about 5 mM, about 0.75 mM to about 1 mM, about 0.75 mM to aboutWSGR Docket No. 52459-726.6011.25 mM, about 0.75 mM to about 1.5 mM, about 0.75 mM to about 1.75 mM, about 0.75 mM to about 2 mM, about 0.75 mM to about 3 mM, about 0.75 mM to about 5 mM, about 1 mM to about 1.25 mM, about 1 mM to about 1.5 mM, about 1 mM to about 1.75 mM, about 1 mM to about 2 mM, about 1 mM to about 3 mM, about 1 mM to about 5 mM, about 1.25 mM to about 1.5 mM, about 1.25 mM to about 1.75 mM, about 1.25 mM to about 2 mM, about 1.25 mM to about 3 mM, about 1.25 mM to about 5 mM, about 1.5 mM to about 1.75 mM, about 1.5 mM to about 2 mM, about 1.5 mM to about 3 mM, about 1.5 mM to about 5 mM, about 1.75 mM to about 2 mM, about 1.75 mM to about 3 mM, about 1.75 mM to about 5 mM, about 2 mM to about 3 mM, about 2 mM to about 5 mM, or about 3 mM to about 5 mM.

[0190] In some embodiments, the concentration (e.g., final concentration) of nonoxynol-9 in the excipient is at least about at least about 0.001% v / v, at least about 0.005% v / v, at least about 0.01% v / v, at least about 0.05% v / v, at least about 0.1% v / v, at least about 0.5% v / v, at least about 1% v / v, at least about 2% v / v, at least about 3% v / v, at least about 4% v / v, at least about 5% v / v, or greater than about 5% v / v. In some embodiments, the concentration (e.g., final concentration) of nonoxynol-9 in the excipient is at most about 5% v / v, at most about 4% v / v, at most about 3% v / v, at most about 2% v / v, at most about 1% v / v, at most about 0.5% v / v, at most about 0.1% v / v, at most about 0.05% v / v, at most about 0.01% v / v, at most about 0.005% v / v, at most about 0.001% v / v, or less than about 0.001% v / v.

[0191] In some embodiments, the concentration (e.g., final concentration) of nonoxynol-9 in the excipient can be between about 0.01 % v / v to about 5 % v / v. In some embodiments, the concentration (e.g., final concentration) of nonoxynol-9 in the excipient can be between at most about 5 % v / v. In some embodiments, the concentration (e.g., final concentration) of nonoxynol- 9 in the excipient can be between about 0.01 % v / v to about 0.05 % v / v, about 0.01 % v / v to about 0.1 % v / v, about 0.01 % v / v to about 0.5 % v / v, about 0.01 % v / v to about 0.75 % v / v, about 0.01 % v / v to about 1 % v / v, about 0.01 % v / v to about 1.25 % v / v, about 0.01 % v / v to about 1.5 % v / v, about 0.01 % v / v to about 1.75 % v / v, about 0.01 % v / v to about 2 % v / v, about 0.01 % v / v to about 3 % v / v, about 0.01 % v / v to about 5 % v / v, about 0.05 % v / v to about 0.1 % v / v, about 0.05 % v / v to about 0.5 % v / v, about 0.05 % v / v to about 0.75 % v / v, about 0.05 % v / v to about 1 % v / v, about 0.05 % v / v to about 1.25 % v / v, about 0.05 % v / v to about 1.5 % v / v, about 0.05 % v / v to about 1.75 % v / v, about 0.05 % v / v to about 2 % v / v, about 0.05 % v / v to about 3 % v / v, about 0.05 % v / v to about 5 % v / v, about 0.1 % v / v to about 0.5 % v / v, about 0.1 % v / v to about 0.75 % v / v, about 0.1 % v / v to about 1 % v / v, about 0.1 % v / v to about 1.25 % v / v, about 0.1 % v / v to about 1.5 % v / v, about 0.1 % v / v to about 1.75 % v / v, about 0.1 % v / v toWSGR Docket No. 52459-726.601 about 2 % v / v, about 0.1 % v / v to about 3 % v / v, about 0.1 % v / v to about 5 % v / v, about 0.5 % v / v to about 0.75 % v / v, about 0.5 % v / v to about 1 % v / v, about 0.5 % v / v to about 1.25 % v / v, about 0.5 % v / v to about 1.5 % v / v, about 0.5 % v / v to about 1.75 % v / v, about 0.5 % v / v to about 2 % v / v, about 0.5 % v / v to about 3 % v / v, about 0.5 % v / v to about 5 % v / v, about 0.75 % v / v to about 1 % v / v, about 0.75 % v / v to about 1.25 % v / v, about 0.75 % v / v to about 1.5 % v / v, about 0.75 % v / v to about 1.75 % v / v, about 0.75 % v / v to about 2 % v / v, about 0.75 % v / v to about 3 % v / v, about 0.75 % v / v to about 5 % v / v, about 1 % v / v to about 1.25 % v / v, about 1 % v / v to about 1.5 % v / v, about 1 % v / v to about 1.75 % v / v, about 1 % v / v to about 2 % v / v, about 1 % v / v to about 3 % v / v, about 1 % v / v to about 5 % v / v, about 1.25 % v / v to about 1.5 % v / v, about 1.25 % v / v to about 1.75 % v / v, about 1.25 % v / v to about 2 % v / v, about 1.25 % v / v to about 3 % v / v, about 1.25 % v / v to about 5 % v / v, about 1.5 % v / v to about 1.75 % v / v, about 1.5 % v / v to about 2 % v / v, about 1.5 % v / v to about 3 % v / v, about 1.5 % v / v to about 5 % v / v, about 1.75 % v / v to about 2 % v / v, about 1.75 % v / v to about 3 % v / v, about 1.75 % v / v to about 5 % v / v, about 2 % v / v to about 3 % v / v, about 2 % v / v to about 5 % v / v, or about 3 % v / v to about 5 % v / v.

[0192] In some embodiments, the concentration (e.g., final concentration) of trehalose in the excipient is at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, at least about 100 mM, at least about 250 mM, at least about 500 mM, at least about 750 mM, at least about 1000 mM, at least about 2000 mM, at least about 3000 mM, at least about 4000 mM, at least about 5000 mM, or greater than about 5000 mM.

[0193] In some embodiments, the concentration (e.g., final concentration) of trehalose in the excipient is at most about 5000 mM, at most about 4000 mM, at most about 3000 mM, at most about 2000 mM, at most about 1000 mM, at most about 750 mM, at most about 500 mM, at most about 250 mM, at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16WSGR Docket No. 52459-726.601 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at most about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, or at most about 1 mM. In some embodiments, the concentration (e.g., final concentration) of trehalose in the excipient can be between about 0.001 M to about 5 M. In some embodiments, the concentration (e.g., final concentration) of trehalose in the excipient can be between about 0.001 M to about 0.005 M, about 0.001 M to about 0.0075 M, about 0.001 M to about 0.01 M, about 0.001 M to about 0.05 M, about 0.001 M to about 0.075 M, about 0.001 M to about 0.1 M, about 0.001 M to about 0.25 M, about 0.001 M to about 0.5 M, about 0.001 M to about 0.75 M, about 0.001 M to about 1 M, about 0.001 M to about 5 M, about 0.005 M to about 0.0075 M, about 0.005 M to about 0.01 M, about 0.005 M to about 0.05 M, about 0.005 M to about 0.075 M, about 0.005 M to about 0.1 M, about 0.005 M to about 0.25 M, about 0.005 M to about 0.5 M, about 0.005 M to about 0.75 M, about 0.005 M to about 1 M, about 0.005 M to about 5 M, about 0.0075 M to about 0.01 M, about 0.0075 M to about 0.05 M, about 0.0075 M to about 0.075 M, about 0.0075 M to about 0.1 M, about 0.0075 M to about 0.25 M, about 0.0075 M to about 0.5 M, about 0.0075 M to about 0.75 M, about 0.0075 M to about 1 M, about 0.0075 M to about 5 M, about 0.01 M to about 0.05 M, about 0.01 M to about 0.075 M, about 0.01 M to about 0.1 M, about 0.01 M to about 0.25 M, about 0.01 M to about 0.5 M, about 0.01 M to about 0.75 M, about 0.01 M to about 1 M, about 0.01 M to about 5 M, about 0.05 M to about 0.075 M, about 0.05 M to about 0.1 M, about 0.05 M to about 0.25 M, about 0.05 M to about 0.5 M, about 0.05 M to about 0.75 M, about 0.05 M to about 1 M, about 0.05 M to about 5 M, about 0.075 M to about 0.1 M, about 0.075 M to about 0.25 M, about 0.075 M to about 0.5 M, about 0.075 M to about 0.75 M, about 0.075 M to about 1 M, about 0.075 M to about 5 M, about 0.1 M to about 0.25 M, about 0.1 M to about 0.5 M, about 0.1 M to about 0.75 M, about 0.1 M to about 1 M, about 0.1 M to about 5 M, about 0.25 M to about 0.5 M, about 0.25 M to about 0.75 M, about 0.25 M to about 1 M, about 0.25 M to about 5 M, about 0.5 M to about 0.75 M, about 0.5 M to about 1 M, about 0.5 M to about 5 M, about 0.75 M to about 1 M, about 0.75 M to about 5 M, or about 1 M to about 5 M.

[0194] In some embodiments, the concentration (e.g., final concentration) of dextran in the excipient is at least about at least about 0.001% w / v, at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.05% w / v, at least about 0.1% w / v, at least about 0.5% w / v, at least about 1% w / v, at least about 2% w / v, at least about 3% w / v, at least about 4% w / v, at least about 5% w / v, at least about 7.5% w / v, at least about 10% w / v, at least about 15% w / v, or greater thanWSGR Docket No. 52459-726.601 about 15% w / v. In some embodiments, the concentration (e.g., final concentration) of dextran in the excipient is at most about 15% w / v, at most about 10% w / v, at most about 7.5% w / v, at most about 5% w / v, at most about 4% w / v, at most about 3% w / v, at most about 2% w / v, at most about 1% w / v, at most about 0.5% w / v, at most about 0.1% w / v, at most about 0.05% w / v, at most about 0.01% w / v, at most about 0.005% w / v, at most about 0.001% w / v, or less than about 0.001% w / v.

[0195] In some embodiments, the concentration (e.g., final concentration) of dextran in the excipient can be between about 0.1 % w / v to about 8 % w / v. In some embodiments, the concentration (e.g., final concentration) of dextran in the excipient can be between at most about 8 % w / v. In some embodiments, the concentration (e.g., final concentration) of dextran in the excipient can be between about 0.1 % w / v to about 0.5 % w / v, about 0.1 % w / v to about 1 % w / v, about 0.1 % w / v to about 1.5 % w / v, about 0.1 % w / v to about 2 % w / v, about 0.1 % w / v to about 2.5 % w / v, about 0.1 % w / v to about 3 % w / v, about 0.1 % w / v to about 3.5 % w / v, about 0.1 % w / v to about 4 % w / v, about 0.1 % w / v to about 5 % w / v, about 0.1 % w / v to about 6 % w / v, about 0.1 % w / v to about 8 % w / v, about 0.5 % w / v to about 1 % w / v, about 0.5 % w / v to about 1.5 % w / v, about 0.5 % w / v to about 2 % w / v, about 0.5 % w / v to about 2.5 % w / v, about 0.5 % w / v to about 3 % w / v, about 0.5 % w / v to about 3.5 % w / v, about 0.5 % w / v to about 4 % w / v, about 0.5 % w / v to about 5 % w / v, about 0.5 % w / v to about 6 % w / v, about 0.5 % w / v to about 8 % w / v, about 1 % w / v to about 1.5 % w / v, about 1 % w / v to about 2 % w / v, about 1 % w / v to about 2.5 % w / v, about 1 % w / v to about 3 % w / v, about 1 % w / v to about 3.5 % w / v, about 1 % w / v to about 4 % w / v, about 1 % w / v to about 5 % w / v, about 1 % w / v to about 6 % w / v, about 1 % w / v to about 8 % w / v, about 1.5 % w / v to about 2 % w / v, about 1.5 % w / v to about 2.5 % w / v, about 1.5 % w / v to about 3 % w / v, about 1.5 % w / v to about 3.5 % w / v, about 1.5 % w / v to about 4 % w / v, about 1.5 % w / v to about 5 % w / v, about 1.5 % w / v to about 6 % w / v, about 1.5 % w / v to about 8 % w / v, about 2 % w / v to about 2.5 % w / v, about 2 % w / v to about 3 % w / v, about 2 % w / v to about 3.5 % w / v, about 2 % w / v to about 4 % w / v, about 2 % w / v to about 5 % w / v, about 2 % w / v to about 6 % w / v, about 2 % w / v to about 8 % w / v, about2.5 % w / v to about 3 % w / v, about 2.5 % w / v to about 3.5 % w / v, about 2.5 % w / v to about 4 % w / v, about 2.5 % w / v to about 5 % w / v, about 2.5 % w / v to about 6 % w / v, about 2.5 % w / v to about 8 % w / v, about 3 % w / v to about 3.5 % w / v, about 3 % w / v to about 4 % w / v, about 3 % w / v to about 5 % w / v, about 3 % w / v to about 6 % w / v, about 3 % w / v to about 8 % w / v, about3.5 % w / v to about 4 % w / v, about 3.5 % w / v to about 5 % w / v, about 3.5 % w / v to about 6 % w / v, about 3.5 % w / v to about 8 % w / v, about 4 % w / v to about 5 % w / v, about 4 % w / v toWSGR Docket No. 52459-726.601 about 6 % w / v, about 4 % w / v to about 8 % w / v, about 5 % w / v to about 6 % w / v, about 5 % w / v to about 8 % w / v, or about 6 % w / v to about 8 % w / v.

[0196] In some embodiments, the concentration (e.g., final concentration) of poly sucrose 400 in the excipient is at least about at least about 0.001% w / v, at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.05% w / v, at least about 0.1% w / v, at least about 0.5% w / v, at least about 1% w / v, at least about 2% w / v, at least about 3% w / v, at least about 4% w / v, at least about 5% w / v, at least about 7.5% w / v, at least about 10% w / v, at least about 15% w / v, or greater than about 15% w / v (g of solute / 100 mL of solution). In some embodiments, the concentration (e.g., final concentration) of polysucrose 400 in the excipient is at most about 15% w / v, at most about 10% w / v, at most about 7.5% w / v, at most about 5% w / v, at most about 4% w / v, at most about 3% w / v, at most about 2% w / v, at most about 1% w / v, at most about 0.5% w / v, at most about 0.1% w / v, at most about 0.05% w / v, at most about 0.01% w / v, at most about 0.005% w / v, at most about 0.001% w / v, or less than about 0.001% w / v (g of solute / 100 mL of solution).

[0197] In some embodiments, the concentration (e.g., final concentration) of polysucrose 400 in the excipient can be between about 0.001 % w / v to about 5 % w / v. In some embodiments, the concentration (e.g., final concentration) of polysucrose 400 in the excipient can be between at most about 5 % w / v. In some embodiments, the concentration (e.g., final concentration) of polysucrose 400 in the excipient can be between about 0.001 % w / v to about 0.01 % w / v, about 0.001 % w / v to about 0.1 % w / v, about 0.001 % w / v to about 0.2 % w / v, about 0.001 % w / v to about 0.3 % w / v, about 0.001 % w / v to about 0.4 % w / v, about 0.001 % w / v to about 0.5 % w / v, about 0.001 % w / v to about 0.75 % w / v, about 0.001 % w / v to about 1 % w / v, about 0.001 % w / v to about 2 % w / v, about 0.001 % w / v to about 3 % w / v, about 0.001 % w / v to about 5 % w / v, about 0.01 % w / v to about 0.1 % w / v, about 0.01 % w / v to about 0.2 % w / v, about 0.01 % w / v to about 0.3 % w / v, about 0.01 % w / v to about 0.4 % w / v, about 0.01 % w / v to about 0.5 % w / v, about 0.01 % w / v to about 0.75 % w / v, about 0.01 % w / v to about 1 % w / v, about 0.01 % w / v to about 2 % w / v, about 0.01 % w / v to about 3 % w / v, about 0.01 % w / v to about 5 % w / v, about 0.1 % w / v to about 0.2 % w / v, about 0.1 % w / v to about 0.3 % w / v, about 0.1 % w / v to about 0.4 % w / v, about 0.1 % w / v to about 0.5 % w / v, about 0.1 % w / v to about 0.75 % w / v, about 0.1 % w / v to about 1 % w / v, about 0.1 % w / v to about 2 % w / v, about 0.1 % w / v to about 3 % w / v, about 0.1 % w / v to about 5 % w / v, about 0.2 % w / v to about 0.3 % w / v, about 0.2 % w / v to about 0.4 % w / v, about 0.2 % w / v to about 0.5 % w / v, about 0.2 % w / v to about 0.75 % w / v, about 0.2 % w / v to about 1 % w / v, about 0.2 % w / v to about 2 % w / v, about 0.2 % w / v toWSGR Docket No. 52459-726.601 about 3 % w / v, about 0.2 % w / v to about 5 % w / v, about 0.3 % w / v to about 0.4 % w / v, about 0.3 % w / v to about 0.5 % w / v, about 0.3 % w / v to about 0.75 % w / v, about 0.3 % w / v to about 1 % w / v, about 0.3 % w / v to about 2 % w / v, about 0.3 % w / v to about 3 % w / v, about 0.3 % w / v to about 5 % w / v, about 0.4 % w / v to about 0.5 % w / v, about 0.4 % w / v to about 0.75 % w / v, about 0.4 % w / v to about 1 % w / v, about 0.4 % w / v to about 2 % w / v, about 0.4 % w / v to about 3 % w / v, about 0.4 % w / v to about 5 % w / v, about 0.5 % w / v to about 0.75 % w / v, about 0.5 % w / v to about 1 % w / v, about 0.5 % w / v to about 2 % w / v, about 0.5 % w / v to about 3 % w / v, about 0.5 % w / v to about 5 % w / v, about 0.75 % w / v to about 1 % w / v, about 0.75 % w / v to about 2 % w / v, about 0.75 % w / v to about 3 % w / v, about 0.75 % w / v to about 5 % w / v, about 1 % w / v to about 2 % w / v, about 1 % w / v to about 3 % w / v, about 1 % w / v to about 5 % w / v, about 2 % w / v to about 3 % w / v, about 2 % w / v to about 5 % w / v, or about 3 % w / v to about 5 % w / v.

[0198] In some embodiments, the concentration (e.g., final concentration) of cyclodextrin in the excipient is at least about 1 mM, at least about 2 mM, at least about 3 mM, at least about 4 mM, at least about 5 mM, at least about 6 mM, at least about 7 mM, at least about 8 mM, at least about 9 mM, at least about 10 mM, at least about 11 mM, at least about 12 mM, at least about 13 mM, at least about 14 mM, at least about 15 mM, at least about 16 mM, at least about 17 mM, at least about 18 mM, at least about 19 mM, at least about 20 mM, at least about 25 mM, at least about 30 mM, at least about 35 mM, at least about 40 mM, at least about 45 mM, at least about 50 mM, at least about 60 mM, at least about 70 mM, at least about 80 mM, at least about 90 mM, at least about 100 mM, at least about 250 mM, at least about 500 mM, at least about 750 mM, at least about 1000 mM, at least about 2000 mM, at least about 3000 mM, at least about 4000 mM, at least about 5000 mM, or greater than about 5000 mM.

[0199] In some embodiments, the concentration (e.g., final concentration) of cyclodextrin in the excipient is at most about 5000 mM, at most about 4000 mM, at most about 3000 mM, at most about 2000 mM, at most about 1000 mM, at most about 750 mM, at most about 500 mM, at most about 250 mM, at most about 100 mM, at most about 90 mM, at most about 80 mM, at most about 70 mM, at most about 60 mM, at most about 50 mM, at most about 45 mM, at most about 40 mM, at most about 35 mM, at most about 30 mM, at most about 25 mM, at most about 20 mM, at most about 19 mM, at most about 18 mM, at most about 17 mM, at most about 16 mM, at most about 15 mM, at most about 14 mM, at most about 13 mM, at most about 12 mM, at most about 11 mM, at most about 10 mM, at most about 9 mM, at most about 8 mM, at mostWSGR Docket No. 52459-726.601 about 7 mM, at most about 6 mM, at most about 5 mM, at most about 4 mM, at most about 3 mM, at most about 2 mM, or at most about 1 mM.

[0200] In some embodiments, the concentration (e.g., final concentration) of cyclodextrin in the excipient can be between about 0.001 M to about 5 M. In some embodiments, the concentration (e.g., final concentration) of cyclodextrin in the excipient can be between at most about 5 M. In some embodiments, the concentration (e.g., final concentration) of cyclodextrin in the excipient can be between about 0.001 M to about 0.005 M, about 0.001 M to about 0.01 M, about 0.001 M to about 0.02 M, about 0.001 M to about 0.03 M, about 0.001 M to about 0.04 M, about 0.001 M to about 0.05 M, about 0.001 M to about 0.1 M, about 0.001 M to about 0.5 M, about 0.001 M to about 1 M, about 0.001 M to about 3 M, about 0.001 M to about 5 M, about 0.005 M to about 0.01 M, about 0.005 M to about 0.02 M, about 0.005 M to about 0.03 M, about 0.005 M to about 0.04 M, about 0.005 M to about 0.05 M, about 0.005 M to about 0.1 M, about 0.005 M to about 0.5 M, about 0.005 M to about 1 M, about 0.005 M to about 3 M, about 0.005 M to about 5 M, about 0.01 M to about 0.02 M, about 0.01 M to about 0.03 M, about 0.01 M to about 0.04 M, about 0.01 M to about 0.05 M, about 0.01 M to about 0.1 M, about 0.01 M to about 0.5 M, about 0.01 M to about 1 M, about 0.01 M to about 3 M, about 0.01 M to about 5 M, about 0.02 M to about 0.03 M, about 0.02 M to about 0.04 M, about 0.02 M to about 0.05 M, about 0.02 M to about 0.1 M, about 0.02 M to about 0.5 M, about 0.02 M to about 1 M, about 0.02 M to about 3 M, about 0.02 M to about 5 M, about 0.03 M to about 0.04 M, about 0.03 M to about 0.05 M, about 0.03 M to about 0.1 M, about 0.03 M to about 0.5 M, about 0.03 M to about 1 M, about 0.03 M to about 3 M, about 0.03 M to about 5 M, about 0.04 M to about 0.05 M, about 0.04 M to about 0.1 M, about 0.04 M to about 0.5 M, about 0.04 M to about 1 M, about 0.04 M to about 3 M, about 0.04 M to about 5 M, about 0.05 M to about 0.1 M, about 0.05 M to about 0.5 M, about 0.05 M to about 1 M, about 0.05 M to about 3 M, about 0.05 M to about 5 M, about 0.1 M to about 0.5 M, about 0.1 M to about 1 M, about 0.1 M to about 3 M, about 0.1 M to about 5 M, about 0.5 M to about 1 M, about 0.5 M to about 3 M, about 0.5 M to about 5 M, about 1 M to about 3 M, about 1 M to about 5 M, or about 3 M to about 5 M.

[0201] The excipient may comprise at least one additional reagent (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or more additional reagents). In some embodiments, the additional reagent can comprise a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof. The additional reagent of the excipient may be configured to stabilize an enzyme. The additional reagent may lower a Cq value of a nucleic acid amplification. In some embodiments, a nucleic acid amplification canWSGR Docket No. 52459-726.601 have a lower Cq value following a composition comprising an excipient described herein, compared to a Cq value of a nucleic acid amplification that does not comprise a composition comprising the excipient.

[0202] A composition described herein may further comprise a sample stabilization buffer. The sample stabilization buffer can comprise one or more reagents. The one or more reagents may be a collapse modifier, a protein stabilizer, a glass transition modifier, or any combination thereof. In some embodiments, the sample stabilization buffer can comprise at least one salt (e.g., 1, 2, 3, 4, 5, or more salts). The sample stabilization buffer may comprise a cyclodextrin, wherein the cyclodextrin can be a cyclodextrin and / or a concentration of a cyclodextrin as described herein. The one or more reagents of the sample stabilization buffer may be optimized for freeze drying. The sample stabilization buffer may be configured to reconstitute a lyophilized sample. Application of the sample stabilization buffer may reconstitute a lyophilized sample and provide for an improved nucleic acid amplification of the sample. In some embodiments, the sample stabilization buffer comprises one or more reducing agents. The one or more reducing agents can be oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, 1 ,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP), or any combination thereof.

[0203] Without wishing to be bound by theory, the composition provided herein may stabilize nucleic acids during the nucleic acid amplification which may improve the precision and / or efficiency of the amplification.

[0204] In some embodiments, a composition (e.g., a recovery buffer) described herein may comprise one or more cucurbituril. Cucurbiturils are macrocyclic molecules made of glycoluril monomers linked by methylene bridges. Cucurbiturils may form host guest complexes in a composition described herein. Without wishing to be bound by theory, cucurbiturils may be advantageous in a sample preparation as they host one or more inhibitory substances in samples (e.g., PAX samples) so that the samples may be run directly without purification. The PAX sample, as used herein, refers to a sample collected in a PAXgene® tube (or PAX tube as used here). The sample collected in the PAXgene® tube can be directly used for further sample processing or subject to amplification reactions using the amplification buffers described herein. The PAXgene® Blood RNA tube comprises one or more solutions configured to stabilize intracellular RNA. In some cases, the PAXgene® Blood RNA tube can comprise tetradecyl trimethyl -ammonium oxalate and / or tartaric acid. To use the PAXgene® Blood RNA tube, the sample may undergo centrifugation. Use of the compositions and / or methods described hereinWSGR Docket No. 52459-726.601 comprising one or more of the buffers described herein may remove the need to centrifuge a sample for sample processing, sample stabilization, and / or sample amplification. In some embodiments, the compositions and / or methods described herein comprising one or more of the buffers described herein can improve stabilization of genetic material (e.g., RNA) compared to that of the PAXgene® Blood RNA system. The use of the compositions and / or methods described herein may enhance stabilization of genetic material (e.g., RNA) to improve quality of an amplified genetic product, reduce time to produce an amplified product, or any combination thereof.

[0205] Cucurbiturils may comprise one or more glycoluril units (e.g., glycoluril monomers). In some embodiments, a cucurbituril of a composition (e.g., recovery buffer) described herein may comprise at least about 1 glycoluril unit, at least about 2 glycoluril units, at least about 3 glycoluril units, at least about 4 glycoluril units, at least about 5 glycoluril units, at least about 6 glycoluril units, at least about 7 glycoluril units, at least about 8 glycoluril units, at least about 9 glycoluril units, at least about 10 glycoluril units, or greater than about 10 glycoluril units. The cucurbituril may be noted as cucurbit[n]uril, where n is an integer designating the number of glycoluril units. In some embodiments, the composition (e.g., a recovery buffer) described herein may comprise cucurbit[l]uril, cucurbit[2]uril, cucurbit[3]uril, cucurbit[4]uril, cucurbit[5]uril, cucurbit[6]uril, cucurbit[7]uril, cucurbit[8]uril, cucurbit[9]uril, or cucurbit

[0010] uril.

[0206] In some aspects, provided herein is a composition for sample amplification. The composition for sample amplification may comprise a nonionic surfactant, a cyclodextrin, a sucrose / epichlorohydrin polymer, or any combination thereof. The composition may be configured to increase a rate of amplification. The amplification may be a nucleic acid amplification (e.g., a PCR or an isothermal nucleic amplification).

[0207] For example, a composition for sample amplification can comprise: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer, wherein the composition is configured to increase a rate of amplification during a nucleic acid amplification.

[0208] The composition for sample amplification may be configured to stabilize one or more enzyme (e.g., a thermostable enzyme). The enzyme can be stabilized during an amplification (e.g., a nucleic acid amplification). The enzyme can be a polymerase, an endonuclease, or a reverse transcriptase, or any combination thereof. In some embodiments, the reverse transcriptase can be an avian myeloblastosis virus (AMV) reverse transcriptase or a murineWSGR Docket No. 52459-726.601 leukemia virus (MMLV) reverse transcriptase. The nonionic surfactant of the composition for sample amplification may be nonoxynol-9.

[0209] The composition for sample amplification may comprise a cyclodextrin as described herein. The composition for sample amplification may comprise a concentration of cyclodextrin. In some embodiments, the cyclodextrin is present at a final concentration in the presence of the sample. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the composition in the presence of the sample is at least about 0.05 mM, at least about 0.1 mM, at least about 0.5 mM, at least about 1.0 mM, at least about 5.0 mM, at least about 10.0 mM, at least about 15.0 mM, at least about 20.0 mM, at least about 25.0 mM, at least about 30.0 mM, at least about 35.0 mM, at least about 40.0 mM, at least about 50.0 mM, at least about 55.0 mM, at least about 60.0 mM, at least about 65.0 mM, at least about 70.0 mM, at least about 75.0 mM, at least about 80.0 mM, at least about 85.0 mM, at least about 90.0 mM, at least about 95.0 mM, at least about 100.0 mM, at least about 500 mM, at least about 1000 mM, at least about 2500 mM, at least about 5000 mM, at least about 7500 mM, at least about 10000 mM, at least about 20000 mM, or greater than about 20000 mM. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the composition in the presence of the sample is at most about 20000 mM, at most about 10000 mM, at most about 7500 mM, at most about 5000 mM, at most about 2500 mM, at most about 1000 mM, at most about 500 mM, at most about 100.0 mM, at most about 95.0 mM, at most about 90.0 mM, at most about 85.0 mM, at most about 80.0 mM, at most about 75.0 mM, at most about 70.0 mM, at most about 65.0 mM, at most about 60.0 mM, at most about 55.0 mM, at most about 50.0 mM, at most about 45.0 mM, at most about 40.0 mM, at most about 35.0 mM, at most about 30.0 mM, at most about 25.0 mM, at most about 20.0 mM, at most about 15.0 mM, at most about 10.0 mM, at most about 5.0 mM, at most about 1.0 mM, at most about 0.5 mM, at most about 0.1 mM, or at most about 0.05 mM.

[0210] In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the composition in the presence of the sample is at least about 0.001% v / v, at least about 0.005% v / v, at least about 0.01% v / v, at least about 0.05% v / v, at least about 0.1% v / v, at least about 0.5% v / v, at least about 1.0% v / v, at least about 1.5% v / v, at least about 2.0% v / v, at least about 3.0% v / v, at least about 4.0% v / v, at least about 5.0% v / v, or greater than about 5.0% v / v. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the composition in the presence of the sample is at most about 5.0% v / v, at most about 4.0% v / v, at most about 3.0% v / v, at most about 2.0% v / v, at most about 1.5% v / v, at most about 1.0% v / v, atWSGR Docket No. 52459-726.601 most about 0.5% v / v, at most about 0.1% v / v, at most about 0.05% v / v, at most about 0.01% v / v, at most about 0.005% v / v, at most about 0.001% v / v, or less than about 0.001% v / v.

[0211] The sucrose / epichlorohydrin polymer of the composition may be poly sucrose 400. In some embodiments, the concentration (e.g., final concentration) of poly sucrose 400 in the composition is at least about 0.001% w / v, at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.05% w / v, at least about 0.1% w / v, at least about 0.5% w / v, at least about 1% w / v, at least about 2% w / v, at least about 3% w / v, at least about 4% w / v, at least about 5% w / v, at least about 7.5% w / v, at least about 10% w / v, at least about 15% w / v, or greater than about 15% w / v (g of solute / 100 mL of solution). In some embodiments, the concentration (e.g., final concentration) of polysucrose 400 in the composition is at most about 15% w / v, at most about 10% w / v, at most about 7.5% w / v, at most about 5% w / v, at most about 4% w / v, at most about 3% w / v, at most about 2% w / v, at most about 1% w / v, at most about 0.5% w / v, at most about 0.1% w / v, at most about 0.05% w / v, at most about 0.01% w / v, at most about 0.005% w / v, at most about 0.001% w / v, or less than about 0.001% w / v (g of solute / 100 mL of solution).

[0212] The composition for sample amplification can comprise one or more salts (e.g., 1, 2, 3, 4, 5, or more salts. The salt may be sodium chloride, potassium chloride, potassium phosphate, or any combination thereof. A final concentration of at least one salt in the composition in the presence of a sample may be at least about 0.0001 M, at least about 0.001 M, at least about 0.005 M, at least about 0.01 M, at least about 0.05 M, at least about 0.1 M, at least about 0.5 M, at least about 1.0 M, at least about 2.0 M, at least about 3.0 M, at least about 4.0 M, at least about 5.0 M, at least about 6.0 M, at least about 7.0 M, at least about 8.0 M, at least about 9.0 M, at least about 10 M, at least about 15 M, or greater than about 15 M. A final concentration of at least one salt in the composition in the presence of a sample may be at most about 15 M, at most about 10 M, at most about 9.0 M, at most about 8.0 M, at most about 7.0 M, at most about 6.0 M, at most about 5.0 M, at most about 4.0 M, at most about 3.0 M, at most about 2.0 M, at most about 1.0 M, at most about 0.5 M, at most about 0.1 M, at most about 0.05 M, at most about 0.01 M, at most about 0.005 M, at most about 0.001 M, at most about 0.0001 M, or less than about 0.0001 M.

[0213] In some embodiments, the composition can comprise an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)- 1,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, aWSGR Docket No. 52459-726.601 thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), tetrahydropyran (THP), or any combination thereof. The composition can comprise a concentration of EDTA and / or Tris as described herein. The composition for sample amplification may comprise one or more agents capable of reducing a disulfide bond (e.g., dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), 2-mercaptoehtanol ( ME), or any combination thereof).

[0214] The composition can comprise one or more sugars and / or sugar alcohols. The one or more sugars and / or sugar alcohols can comprise sucrose, lactose, trehalose, dextran, erythritol, arabitol, xylitol, sorbitol, mannitol, or any combination thereof. In some embodiments, a composition described herein may comprise a final concentration of sugar and / or sugar alcohol of at least about 0.0001 M, at least about 0.001 M, at least about 0.005 M, at least about 0.01 M, at least about 0.05 M, at least about 0.1 M, at least about 0.5 M, at least about 1.0 M, at least about 2.0 M, at least about 3.0 M, at least about 4.0 M, at least about 5.0 M, at least about 6.0 M, at least about 7.0 M, at least about 8.0 M, at least about 9.0 M, at least about 10 M, at least about 15 M, or greater than about 15 M. In some embodiments, a composition described herein may comprise a final concentration of sugar and / or sugar alcohol of at most about 15 M, at most about 10 M, at most about 9.0 M, at most about 8.0 M, at most about 7.0 M, at most about 6.0 M, at most about 5.0 M, at most about 4.0 M, at most about 3.0 M, at most about 2.0 M, at most about 1.0 M, at most about 0.5 M, at most about 0.1 M, at most about 0.05 M, at most about 0.01 M, at most about 0.005 M, at most about 0.001 M, at most about 0.0001 M, or less than about 0.0001 M.

[0215] In some embodiments, a composition described herein may comprise a final concentration of sugar and / or sugar alcohol of at least about 0.001% w / v, at least about 0.005% w / v, at least about 0.01% w / v, at least about 0.05% w / v, at least about 0.1% w / v, at least about 0.5% w / v, at least about 1% w / v, at least about 2% w / v, at least about 3% w / v, at least about 4% w / v, at least about 5% w / v, at least about 7.5% w / v, at least about 10% w / v, at least about 15% w / v, or greater than about 15% w / v (g of solute / 100 mL of solution). In some embodiments, a composition described herein may comprise a final concentration of sugar and / or sugar alcohol of at most about 15% w / v, at most about 10% w / v, at most about 7.5% w / v, at most about 5% w / v, at most about 4% w / v, at most about 3% w / v, at most about 2% w / v, at most about 1% w / v, at most about 0.5% w / v, at most about 0.1% w / v, at most about 0.05% w / v, at most about 0.01% w / v, at most about 0.005% w / v, at most about 0.001% w / v, or less than about 0.001% w / v (g of solute / 100 mL of solution).WSGR Docket No. 52459-726.601

[0216] The composition for sample amplification may comprise at least one additional reagent (e.g., 1, 2, 3, 4, 5, 6, 7, 8, or more additional reagents). In some embodiments, the additional reagent can comprise a base, Brij 98, guanidinium thiocyanate (GITC), methionine, nondetergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof. A composition described herein may be lyophilized. The composition for sample amplification may be configured to stabilize enzymatic activity of a thermostable enzyme for use during the nucleic acid amplification. The thermostable enzyme may be a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, a IsoFast™ Bst, any mutants thereof, or any combination thereof.

[0217] In some aspects, provided herein is a composition comprising a sample processing buffer. The sample processing buffer may comprise one or more of a detergent, a solubilizer, or a cyclodextrin. The composition may comprise a sample amplification buffer. The sample amplification buffer may comprise one or more of a nonionic surfactant, a cyclodextrin, or a sucrose / epichlorohydrin polymer. The composition may comprise a sample stabilization buffer. The sample stabilization buffer may be configured to stabilize one or more enzymes. The enzyme may be in a nucleic acid amplification.

[0218] For example, a composition may comprise: a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer; and a sample stabilization buffer configured to stabilize an enzyme in a nucleic acid amplification.

[0219] The solubilizer and cyclodextrin of the composition may be configured to shorten a cycle threshold (Ct) value and / or a time to result value. In some cases, quantification cycle Cq value is also used and it can be used interchangeably with Ct value. In some embodiments, the solubilizer and cyclodextrin of the composition may be configured to shorten a cycle threshold (Ct) value and / or a time to result value of a nucleic acid amplification compared to a cycle threshold (Ct) value and / or a time to result value in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually. In some embodiments, the solubilizer and / or cyclodextrin described herein are configured to shorten a cycle threshold value to at most about 60, at most about 50, at most about 40, at most about 30, at most about 25, at most about 20, at most about 19, at most about 18, at most about 17, at most about 16, at most about 15, at most about 14, at most about 13, at most about 12, at most aboutWSGR Docket No. 52459-726.60111, at most about 10, at most about 9, at most about 8, at most about 7, at most about 6, at most about 5, at most about 4, at most about 3, at most about 2, or at most about 1. In some embodiments, the solubilizer and / or cyclodextrin described herein are configured to shorten a time to result value to at most about 15 minutes, at most about 14 minutes, at most about 13 minutes, at most about 12 minutes, at most about 11 minutes, at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6 minutes, at most about 5 minutes or less.

[0220] The detergent may be part of a lysis buffer, wherein the lysis buffer can comprise a chelating agent. The chelating agent can be deferiprone, ethylenediamine, 1,10-Phenanthroline, oxalic acid, pentetic acid, deferasirox, deferoxamine, deferoxamine mesylate, N,N,N',N'- tetrakis(2-pyridinylmethyl)-l,2-ethanediamine (TPEN), or any combination thereof. The composition can comprise a reducing agent. The reducing agent may be oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, l,2-bis(o- aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), tetrahydropyran (THP), or any combination thereof. The lysis buffer can comprise any compounds (e.g., components) as described herein. The composition can comprise a final concentration of EGTA, EDTA, TCEP, and / or Tris in a lysis buffer in the presence of a sample. The final concentration of EGTA, EDTA, TCEP, and / or Tris may be a concentration as described herein.

[0221] The concentration of Tris in a recovery buffer described herein may be greater than a concentration of Tris after mixed with a sample. For example, a concentration of Tris in a recovery buffer described herein may be at least about 100 mM, at least about 200 mM, at least about 300 mM, at least about 400 mM, at least about 500 mM, at least about 600 mM, at least about 700 mM, at least about 800 mM, at least about 900 mM, at least about 1000 mM, at least about 1500 mM, at least about 2000 mM, or greater than about 2000 mM. A concentration of Tris in a recovery buffer described herein may be at most about 2000 mM, at most about 1500 mM, at most about 1000 mM, at most about 900 mM, at most about 800 mM, at most about 700 mM, at most about 600 mM, at most about 500 mM, at most about 400 mM, at most about 300 mM, at most about 200 mM, at most about 100 mM, or less than about 100 mM.

[0222] The detergent may be present at a concentration (e.g., a final concentration) that can be sufficient for lysing cells. For example, when mixed with a sample described herein, the detergent of the sample processing buffer can lyse the cells. The cyclodextrin may be present at a concentration (e.g., a final concentration) that can be sufficient for isolating the detergent in the composition. For example, when mixed with a sample described herein, the cyclodextrin ofWSGR Docket No. 52459-726.601 the sample processing buffer can isolate the detergent (e.g., a portion of the detergent) in the composition. The final concentration of the detergent, solubilizer, and / or cyclodextrin can comprise a final concentration as described herein.

[0223] As another example, a composition for sample processing may comprise a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer; a stabilizing agent comprising tetradecyl trimethyl-ammonium oxalate and / or tartaric acid, and wherein the composition is configured to increase a rate of amplification during a nucleic acid amplification.

[0224] As another example, a composition for sample processing may comprise a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; a stabilizing agent comprising tetradecyl trimethyl-ammonium oxalate and / or tartaric acid, and wherein the composition is configured to stabilize an enzyme during a nucleic acid amplification, and wherein the composition is configured to reduce and / or eliminate activity of a degrading nuclease.

[0225] The composition may comprise one or more cucurbituril. The cucurbiturils may comprise one or more glycoluril units (e.g., glycoluril monomers). In some embodiments, a cucurbituril of a composition described herein may comprise at least about 1 glycoluril unit, at least about 2 glycoluril units, at least about 3 glycoluril units, at least about 4 glycoluril units, at least about 5 glycoluril units, at least about 6 glycoluril units, at least about 7 glycoluril units, at least about 8 glycoluril units, at least about 9 glycoluril units, at least about 10 glycoluril units, or greater than about 10 glycoluril units. The cucurbituril may be noted as cucurbit[n]uril, where n is an integer designating the number of glycoluril units. In some embodiments, the composition described herein may comprise cucurbitfl ]uril, cucurbit[2]uril, cucurbit[3]uril, cucurbit[4]uril, cucurbit[5]uril, cucurbit[6]uril, cucurbit[7]uril, cucurbit[8]uril, cucurbit[9]uril, or cucurbitfl 0]uril.

[0226] The composition may further comprise a sample. The sample may be a biological sample. The sample may be a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, and / or a lymph sample, raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof.WSGR Docket No. 52459-726.601

[0227] The blood sample may be obtained from the subject. The blood sample may be obtained at least about 10 seconds, at least about 20 seconds, at least about 30 seconds, at least about 40 seconds, at least about 50 seconds, at least about 1 minute, at least about 5 minutes, at least about 10 minutes, at least about 20 minutes, at least about 30 minutes, at least about 1 hour, at least about 6 hours, at least about 12 hours, at least about 24 hours, or greater than about 24 hours prior to sample processing. The blood sample may be collected in a blood collection tube. In some embodiments, the blood collection tube can comprise a stabilizing agent. The stabilizing agent can stabilize genetic material (e.g., DNA and / or RNA). In some embodiments, the stabilizing agent can comprise tetradecyl trimethyl-ammonium oxalate, tartaric acid, or any combination thereof. In some embodiments, the blood sample may be contacted with the sample processing buffer described herein. The blood sample may be contacted with the sample processing buffer directly. Prior to being contacted by the sample processing buffer, the blood sample may not be subjected to processing (e.g., contacted by another sample processing composition and / or methods). Prior to being contacted by the sample processing buffer, the blood sample may be subjected to processing (e.g., contacted by another sample processing composition and / or methods). In some embodiments, the blood sample may not be processed by centrifugation, spin column, or any combination thereof prior to contacting the sample processing buffer described herein. In some embodiments, the blood sample may be processed by centrifugation, spin column, or any combination thereof prior to contacting the sample processing buffer described herein.Methods for Sample Processing, Stabilization, and Amplification

[0228] In some aspects, the present disclosure provides methods for processing a sample. In some aspects, the method comprises mixing a sample with the lysis buffer described herein. In some embodiments, the method comprises mixing a sample with the recovery buffer described herein. In some embodiments, the method comprises mixing a sample in a mixture of the lysis buffer and recovery buffer described herein.

[0229] In an aspect, the present disclosure provides the method of processing sample comprising (a) contacting the sample with a lysis buffer comprising a detergent, (b) contacting the sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing the sample to generate a processed sample in a mixture comprising the detergent, the solubilizer, and the cyclodextrin. In some cases, the method comprises contacting the lysis buffer in the first step, and then contacting the sample mixed with the lysis buffer with aWSGR Docket No. 52459-726.601 recovery buffer to provide a processed sample. The processed sample can then be contacted with a reaction buffer for amplification. The lysis buffer, the recovery buffer and the amplification buffer can be contacted with the sample at different steps. Separating the lysis buffer and the recovery buffer can offer a flexible system where the processed sample can be compatible with any existing amplification reactions, e.g., PCR or isothermal amplifications.

[0230] In some embodiments, the sample is obtained from a subject prior to the sample processing methods of the present disclosure. In some embodiments, the subject has or is suspected of having a disease, a disorder, a condition, or an infection. In some embodiments, the subject is a non-infectious individual. In some embodiments, the subject is a healthy subject.

[0231] In some embodiment, the sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, and / or a lymph sample for nucleic acid amplification. In some embodiments, the swab sample comprises a vaginal swab, an oral swab, and / or a rectal swab. In some embodiments, the sample comprises raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof. In some embodiments, the sample is a solid sample. In some embodiments, the sample is a liquid sample. In some embodiments, the sample is obtained from a subject. In some embodiments, the subject has a disease, a condition, or an infection. In some embodiments, the sample comprises a biological sample. In some embodiments, the sample comprises a purified sample. In some embodiments, the sample is a combination of two, three, four, five, or more types of samples. In some embodiments, the sample comprises one, two, three, four, five, six, seven, eight, nine, ten, or more target nucleic acid molecules.

[0232] The sample can be contacted with a lysis buffer. The sample can be a liquid sample or a solid sample. In some cases, the sample is a liquid sample and it can be mixed with the lysis buffer with 1 : 10, 1 :20, 1 :50, 1 :60, 1 :70, 1 :80, 1 :90, 1 :100, 1 :200, 1 :500, 1 : 1000 ratio of volume to volume with a lysis buffer such that the final concertation of the lysis buffer is lx lysis buffer. In some cases, the sample can be obtained on a swab and in such cases, the swab can be contacted with lx lysis buffer. The quantity of sample to be mixed with the lysis buffer can depend on the type of sample and / or the amplification method conducted on the sample. In someWSGR Docket No. 52459-726.601 embodiments, the sample is a quantity of liquid mixed with the lysis buffer for a downstream application (e.g., nucleic acid amplification).

[0233] In some embodiments, the quantity of the sample to be mixed with the lysis buffer comprises at least about 0.1 pl, at least about 0.2 pl, at least about 0.3 pl, at least about 0.4 pl, at least about 0.5 pl, at least about 0.6 pl, at least about 0.7 pl, at least about 0.8 pl, at least about 0.9 pl, at least about 1.0 pl, at least about 2.0 pl, at least about 3.0 pl, at least about 4.0 pl, at least about 5.0 pl, at least about 6.0 pl, at least about 7.0 pl, at least about 8.0 pl, at least about 9.0 pl, at least about 10.0 pl, at least about 11.0 pl, at least about 12.0 pl, at least about 13.0 pl, at least about 14.0 pl, at least about 15.0 pl, at least about 16.0 pl, at least about 17.0 pl, at least about 18.0 pl, at least about 19.0 pl, at least about 20.0 pl, at least about 25.0 pl, at least about 30.0 pl, at least about 35.0 pl, at least about 40.0 pl, at least about 45.0 pl, at least about 50.0 pl, at least about 75.0 pl, or at least about 100.0 pl.

[0234] In some embodiments, the quantity of the sample comprises at most about 100.0 pl, at most about 75.0 pl, at most about 50.0 pl, at most about 45.0 pl, at most about 40.0 pl, at most about 35.0 pl, at most about 30.0 pl, at most about 25.0 pl, at most about 20.0 pl, at most about 19.0 pl, at most about 18.0 pl, at most about 17.0 pl, at most about 16.0 pl, at most about 15.0 pl, at most about 14.0 pl, at most about 13.0 pl, at most about 12.0 pl, at most about 11.0 pl, at most about 10.0 pl, at most about 9.0 pl, at most about 8.0 pl, at most about 7.0 pl, at most about 6.0 pl, at most about 5.0 pl, at most about 4.0 pl, at most about 3.0 pl, at most about 2.0 pl, at most about 1.0 pl, at most about 0.9 pl, at most about 0.8 pl, at most about 0.7 pl, at most about 0.6 pl, at most about 0.5 pl, at most about 0.4 pl, at most about 0.3 pl, at most about 0.2 pl, or at most about 0.1 pl.

[0235] In some embodiments, the quantity of the sample comprises about 0.1 pl to about 100 pl. In some embodiments, the quantity of the sample comprises about 0.1 pl to about 0.5 pl, about 0.1 pl to about 0.75 pl, about 0.1 pl to about 1 pl, about 0.1 pl to about 5 pl, about 0.1 pl to about 7.5 pl, about 0.1 pl to about 10 pl, about 0.1 pl to about 20 pl, about 0.1 pl to about 25 pl, about 0.1 pl to about 50 pl, about 0.1 pl to about 75 pl, about 0.1 pl to about 100 pl, about 0.5 pl to about 0.75 pl, about 0.5 pl to about 1 pl, about 0.5 pl to about 5 pl, about 0.5 pl to about 7.5 pl, about 0.5 pl to about 10 pl, about 0.5 pl to about 20 pl, about 0.5 pl to about 25 pl, about 0.5 pl to about 50 pl, about 0.5 pl to about 75 pl, about 0.5 pl to about 100 pl, about 0.75 pl to about 1 pl, about 0.75 pl to about 5 pl, about 0.75 pl to about 7.5 pl, about 0.75 pl to about 10 pl, about 0.75 pl to about 20 pl, about 0.75 pl to about 25 pl, about 0.75 pl to about 50 pl, about 0.75 pl to about 75 pl, about 0.75 pl to about 100 pl, about 1 pl to about 5 pl, about 1 plWSGR Docket No. 52459-726.601 to about 7.5 pl, about 1 pl to about 10 pl, about 1 pl to about 20 pl, about 1 pl to about 25 pl, about 1 pl to about 50 pl, about 1 pl to about 75 pl, about 1 pl to about 100 pl, about 5 pl to about 7.5 pl, about 5 pl to about 10 pl, about 5 pl to about 20 pl, about 5 pl to about 25 pl, about 5 pl to about 50 pl, about 5 pl to about 75 pl, about 5 pl to about 100 pl, about 7.5 pl to about 10 pl, about 7.5 pl to about 20 pl, about 7.5 pl to about 25 pl, about 7.5 pl to about 50 pl, about 7.5 pl to about 75 pl, about 7.5 pl to about 100 pl, about 10 pl to about 20 pl, about 10 pl to about 25 pl, about 10 pl to about 50 pl, about 10 pl to about 75 pl, about 10 pl to about 100 pl, about 20 pl to about 25 pl, about 20 pl to about 50 pl, about 20 pl to about 75 pl, about 20 pl to about 100 pl, about 25 pl to about 50 pl, about 25 pl to about 75 pl, about 25 pl to about 100 pl, about 50 pl to about 75 pl, about 50 pl to about 100 pl, or about 75 pl to about 100 pl.

[0236] In some embodiments, there is a copy number of target nucleic acid molecules to be processed in the sample. In some embodiments, the copy number of target nucleic acid molecules can be at least about 1, at least about 2, at least about 3, at least about 4, at least about5, at least about 6, at least about 7, at least about 8, at least about 9, at least about 10, at least about 20, at least about 30, at least about 40, at least about 50, at least about 75, at least about 100, at least about 150, at least about 200, at least about 250, at least about 300, at least about 350, at least about 400, at least about 450, at least about 500, at least about 750, at least about 1000, at least about 1500, at least about 2000, at least about 2500, at least about 5000, at least about 7500, at least about 10,000, at least about 25,000, at least about 50,000, at least about 75,000, at least about 100,000, at least about 250,000, or at least about 500,000 copies. In some embodiments, the copy number of target nucleic acid molecules can be at most about 500,000, at most about 250,000, at most about 100,000, at most about 75,000, at most about 50,000, at most about 25,000, at most about 10,000, at most about 7500, at most about 5000, at most about 2500, at most about 1000, at most about 750, at most about 500, at most about 450, at most about 400, at most about 350, at most about 300, at most about 250, at most about 200, at most about 150, at most about 100, at most about 75, at most about 50, at most about 40, at most about 30, at most about 20, at most about 10, at most about 9, at most about 8, at most about 7, at most about6, at most about 5, at most about 4, at most about 3, at most about 2, or at most about 1.

[0237] In some embodiments, the copy number of target nucleic acid molecules can be about 1 to about 500,000. In some embodiments, the copy number of target nucleic acid molecules can be about 1 to about 5, about 1 to about 10, about 1 to about 50, about 1 to about 100, about 1 to about 1,000, about 1 to about 2,500, about 1 to about 5,000, about 1 to about 10,000, about 1 to about 50,000, about 1 to about 100,000, about 1 to about 500,000, about 5 to about 10, about 5WSGR Docket No. 52459-726.601 to about 50, about 5 to about 100, about 5 to about 1,000, about 5 to about 2,500, about 5 to about 5,000, about 5 to about 10,000, about 5 to about 50,000, about 5 to about 100,000, about 5 to about 500,000, about 10 to about 50, about 10 to about 100, about 10 to about 1,000, about 10 to about 2,500, about 10 to about 5,000, about 10 to about 10,000, about 10 to about 50,000, about 10 to about 100,000, about 10 to about 500,000, about 50 to about 100, about 50 to about 1,000, about 50 to about 2,500, about 50 to about 5,000, about 50 to about 10,000, about 50 to about 50,000, about 50 to about 100,000, about 50 to about 500,000, about 100 to about 1,000, about 100 to about 2,500, about 100 to about 5,000, about 100 to about 10,000, about 100 to about 50,000, about 100 to about 100,000, about 100 to about 500,000, about 1,000 to about 2,500, about 1,000 to about 5,000, about 1,000 to about 10,000, about 1,000 to about 50,000, about 1,000 to about 100,000, about 1,000 to about 500,000, about 2,500 to about 5,000, about 2,500 to about 10,000, about 2,500 to about 50,000, about 2,500 to about 100,000, about 2,500 to about 500,000, about 5,000 to about 10,000, about 5,000 to about 50,000, about 5,000 to about 100,000, about 5,000 to about 500,000, about 10,000 to about 50,000, about 10,000 to about 100,000, about 10,000 to about 500,000, about 50,000 to about 100,000, about 50,000 to about 500,000, or about 100,000 to about 500,000.

[0238] In some cases, the sample preparation methods and compositions of the present disclosure provide for a greater concentration of target nucleic acid molecules from a processed sample compared to a concentration of target nucleic acid molecules in a sample processed by a different method. In some embodiments, a concentration of one or more different target nucleic acid molecules is at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, or at least about 50%, compared to a concentration of one or more different target nucleic acid molecules of an otherwise identical sample processed by SDS, polysorbate 80, and / or a cyclodextrin individually.

[0239] In some embodiments, the detergent comprises a sodium dodecyl sulfate (SDS). In some embodiments, the detergent comprises sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof. In some embodiments, the detergent is an ionic detergent. In some embodiments, the detergent is a non-ionic detergent.

[0240] In some embodiments, the solubilizer is a non-ionic surfactant. In some embodiments, the solubilizer comprises a polysorbate. The polysorbate may be polyoxyethylene (20) sorbitan monooleate (e.g., polysorbate 80), polyoxyethylene (20) sorbitan monolaurate (e.g., polysorbate 20), polyoxyethylene (20) sorbitan monopalmitate (e.g., polysorbate 40), polyoxyethylene (20)WSGR Docket No. 52459-726.601 sorbitan monostearate (e.g., polysorbate 60), or a functional variant thereof. In some embodiments, the solubilizer is a Tergitol™ surfactant, a Triton™ surfactant, or a Igepal® surfactant. In some embodiments, the solubilizer is an alkoxylate or a cocamide. In some embodiments, the solubilizer is decyl glucoside, alkyl polyglycoside, lauryl glucoside, sorbitan tristearate, or a niosome.

[0241] In some embodiments, the sample is contacted with the lysis buffer and the recovery buffer simultaneously. In some embodiments, the sample is contacted with the lysis buffer and the recovery buffer concurrently in the same mixture. In some embodiments, the sample is submerged in the lysis buffer. In some embodiments, mixing of the sample and lysis buffer is by a vortex and / or by hand. In some embodiments, the sample is not mixed with the lysis buffer.

[0242] In some embodiments, the sample is incubated for a duration of time sufficient for lysis of the cells of the sample. In some embodiments, the sample is incubated for between 1 to 60 minutes in the lysis buffer. In some embodiments, the sample is incubated for at least about 15 seconds, at least about 30 seconds, at least about 1 minute, at least about 1.5 minutes, at least about 2 minutes, at least about 2.5 minutes, at least about 3 minutes, at least about 3.5 minutes, at least about 4 minutes, at least about 4.5 minutes, at least about 5 minutes, at least about 5.5 minutes, at least about 6 minutes, at least about 6.5 minutes, at least about 7 minutes, at least about 8 minutes, at least about 9 minutes, at least about 10 minutes, at least about 12 minutes, at least about 15 minutes, at least about 20 minutes, at least about 25 minutes, at least about 30 minutes, at least about 40 minutes, at least about 50 minutes, at least about 60 minutes, at least about 90 minutes, or at least about 120 minutes in the lysis buffer. In some embodiments, the sample is incubated for at most about 120 minutes, at most about 90 minutes, at most about 60 minutes, at most about 50 minutes, at most about 40 minutes, at most about 30 minutes, at most about 25 minutes, at most about 20 minutes, at most about 15 minutes, at most about 12 minutes, at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6.5 minutes, at most about 6 minutes, at most about 5.5 minutes, at most about 5 minutes, at most about 4.5 minutes, at most about 4 minutes, at most about 3.5 minutes, at most about 3 minutes, at most about 2.5 minutes, at most about 2 minutes, at most about 1.5 minutes, at most about 1 minute, at most about 30 seconds, or at most about 15 seconds in the lysis buffer.

[0243] In some embodiments, the sample is incubated for about 1 minute to about 15 minutes in the lysis buffer. In some embodiments, the sample is incubated for about 1 minute to about 2 minutes, about 1 minute to about 2.5 minutes, about 1 minute to about 3 minutes, about 1 minuteWSGR Docket No. 52459-726.601 to about 3.5 minutes, about 1 minute to about 4 minutes, about 1 minute to about 5 minutes, about 1 minute to about 6 minutes, about 1 minute to about 7 minutes, about 1 minute to about 7.5 minutes, about 1 minute to about 10 minutes, about 1 minute to about 15 minutes, about 2 minutes to about 2.5 minutes, about 2 minutes to about 3 minutes, about 2 minutes to about 3.5 minutes, about 2 minutes to about 4 minutes, about 2 minutes to about 5 minutes, about 2 minutes to about 6 minutes, about 2 minutes to about 7 minutes, about 2 minutes to about 7.5 minutes, about 2 minutes to about 10 minutes, about 2 minutes to about 15 minutes, about 2.5 minutes to about 3 minutes, about 2.5 minutes to about 3.5 minutes, about 2.5 minutes to about 4 minutes, about 2.5 minutes to about 5 minutes, about 2.5 minutes to about 6 minutes, about 2.5 minutes to about 7 minutes, about 2.5 minutes to about 7.5 minutes, about 2.5 minutes to about 10 minutes, about 2.5 minutes to about 15 minutes, about 3 minutes to about 3.5 minutes, about 3 minutes to about 4 minutes, about 3 minutes to about 5 minutes, about 3 minutes to about 6 minutes, about 3 minutes to about 7 minutes, about 3 minutes to about 7.5 minutes, about 3 minutes to about 10 minutes, about 3 minutes to about 15 minutes, about 3.5 minutes to about 4 minutes, about 3.5 minutes to about 5 minutes, about 3.5 minutes to about 6 minutes, about 3.5 minutes to about 7 minutes, about 3.5 minutes to about 7.5 minutes, about 3.5 minutes to about 10 minutes, about 3.5 minutes to about 15 minutes, about 4 minutes to about 5 minutes, about 4 minutes to about 6 minutes, about 4 minutes to about 7 minutes, about 4 minutes to about 7.5 minutes, about 4 minutes to about 10 minutes, about 4 minutes to about 15 minutes, about 5 minutes to about 6 minutes, about 5 minutes to about 7 minutes, about 5 minutes to about 7.5 minutes, about 5 minutes to about 10 minutes, about 5 minutes to about 15 minutes, about 6 minutes to about 7 minutes, about 6 minutes to about 7.5 minutes, about 6 minutes to about 10 minutes, about 6 minutes to about 15 minutes, about 7 minutes to about 7.5 minutes, about 7 minutes to about 10 minutes, about 7 minutes to about 15 minutes, about 7.5 minutes to about 10 minutes, about 7.5 minutes to about 15 minutes, or about 10 minutes to about 15 minutes in the lysis buffer.

[0244] In some embodiments, the sample is incubated for about 1 minute to about 120 minutes. In some embodiments, the sample is incubated for about 1 minute to about 5 minutes, about 1 minute to about 10 minutes, about 1 minute to about 15 minutes, about 1 minute to about 20 minutes, about 1 minute to about 25 minutes, about 1 minute to about 30 minutes, about 1 minute to about 45 minutes, about 1 minute to about 60 minutes, about 1 minute to about 75 minutes, about 1 minute to about 90 minutes, about 1 minute to about 120 minutes, about 5 minutes to about 10 minutes, about 5 minutes to about 15 minutes, about 5 minutes to about 20WSGR Docket No. 52459-726.601 minutes, about 5 minutes to about 25 minutes, about 5 minutes to about 30 minutes, about 5 minutes to about 45 minutes, about 5 minutes to about 60 minutes, about 5 minutes to about 75 minutes, about 5 minutes to about 90 minutes, about 5 minutes to about 120 minutes, about 10 minutes to about 15 minutes, about 10 minutes to about 20 minutes, about 10 minutes to about 25 minutes, about 10 minutes to about 30 minutes, about 10 minutes to about 45 minutes, about 10 minutes to about 60 minutes, about 10 minutes to about 75 minutes, about 10 minutes to about 90 minutes, about 10 minutes to about 120 minutes, about 15 minutes to about 20 minutes, about 15 minutes to about 25 minutes, about 15 minutes to about 30 minutes, about 15 minutes to about 45 minutes, about 15 minutes to about 60 minutes, about 15 minutes to about 75 minutes, about 15 minutes to about 90 minutes, about 15 minutes to about 120 minutes, about 20 minutes to about 25 minutes, about 20 minutes to about 30 minutes, about 20 minutes to about 45 minutes, about 20 minutes to about 60 minutes, about 20 minutes to about 75 minutes, about 20 minutes to about 90 minutes, about 20 minutes to about 120 minutes, about 25 minutes to about 30 minutes, about 25 minutes to about 45 minutes, about 25 minutes to about 60 minutes, about 25 minutes to about 75 minutes, about 25 minutes to about 90 minutes, about 25 minutes to about 120 minutes, about 30 minutes to about 45 minutes, about 30 minutes to about 60 minutes, about 30 minutes to about 75 minutes, about 30 minutes to about 90 minutes, about 30 minutes to about 120 minutes, about 45 minutes to about 60 minutes, about 45 minutes to about 75 minutes, about 45 minutes to about 90 minutes, about 45 minutes to about 120 minutes, about 60 minutes to about 75 minutes, about 60 minutes to about 90 minutes, about 60 minutes to about 120 minutes, about 75 minutes to about 90 minutes, about 75 minutes to about 120 minutes, or about 90 minutes to about 120 minutes.

[0245] In some embodiments, the sample is incubated for about 3 hours to about 48 hours. In some embodiments, the sample is incubated for about 3 hours to about 4 hours, about 3 hours to about 5 hours, about 3 hours to about 10 hours, about 3 hours to about 12 hours, about 3 hours to about 15 hours, about 3 hours to about 18 hours, about 3 hours to about 24 hours, about 3 hours to about 30 hours, about 3 hours to about 36 hours, about 3 hours to about 40 hours, about 3 hours to about 48 hours, about 4 hours to about 5 hours, about 4 hours to about 10 hours, about 4 hours to about 12 hours, about 4 hours to about 15 hours, about 4 hours to about 18 hours, about 4 hours to about 24 hours, about 4 hours to about 30 hours, about 4 hours to about 36 hours, about 4 hours to about 40 hours, about 4 hours to about 48 hours, about 5 hours to about 10 hours, about 5 hours to about 12 hours, about 5 hours to about 15 hours, about 5 hours to about 18 hours, about 5 hours to about 24 hours, about 5 hours to about 30 hours, about 5 hours toWSGR Docket No. 52459-726.601 about 36 hours, about 5 hours to about 40 hours, about 5 hours to about 48 hours, about 10 hours to about 12 hours, about 10 hours to about 15 hours, about 10 hours to about 18 hours, about 10 hours to about 24 hours, about 10 hours to about 30 hours, about 10 hours to about 36 hours, about 10 hours to about 40 hours, about 10 hours to about 48 hours, about 12 hours to about 15 hours, about 12 hours to about 18 hours, about 12 hours to about 24 hours, about 12 hours to about 30 hours, about 12 hours to about 36 hours, about 12 hours to about 40 hours, about 12 hours to about 48 hours, about 15 hours to about 18 hours, about 15 hours to about 24 hours, about 15 hours to about 30 hours, about 15 hours to about 36 hours, about 15 hours to about 40 hours, about 15 hours to about 48 hours, about 18 hours to about 24 hours, about 18 hours to about 30 hours, about 18 hours to about 36 hours, about 18 hours to about 40 hours, about 18 hours to about 48 hours, about 24 hours to about 30 hours, about 24 hours to about 36 hours, about 24 hours to about 40 hours, about 24 hours to about 48 hours, about 30 hours to about 36 hours, about 30 hours to about 40 hours, about 30 hours to about 48 hours, about 36 hours to about 40 hours, about 36 hours to about 48 hours, or about 40 hours to about 48 hours.

[0246] In some embodiments, the sample is incubated in the lysis buffer at room temperature. In some embodiments, the sample at room temperature for about 1 minute to about 15 minutes in the lysis buffer. In some embodiments, the sample is incubated for about 1 minute to about 2 minutes, about 1 minute to about 2.5 minutes, about 1 minute to about 3 minutes, about 1 minute to about 3.5 minutes, about 1 minute to about 4 minutes, about 1 minute to about 5 minutes, about 1 minute to about 6 minutes, about 1 minute to about 7 minutes, about 1 minute to about 7.5 minutes, about 1 minute to about 10 minutes, about 1 minute to about 15 minutes, about 2 minutes to about 2.5 minutes, about 2 minutes to about 3 minutes, about 2 minutes to about 3.5 minutes, about 2 minutes to about 4 minutes, about 2 minutes to about 5 minutes, about 2 minutes to about 6 minutes, about 2 minutes to about 7 minutes, about 2 minutes to about 7.5 minutes, about 2 minutes to about 10 minutes, about 2 minutes to about 15 minutes, about 2.5 minutes to about 3 minutes, about 2.5 minutes to about 3.5 minutes, about 2.5 minutes to about 4 minutes, about 2.5 minutes to about 5 minutes, about 2.5 minutes to about 6 minutes, about 2.5 minutes to about 7 minutes, about 2.5 minutes to about 7.5 minutes, about 2.5 minutes to about 10 minutes, about 2.5 minutes to about 15 minutes, about 3 minutes to about 3.5 minutes, about 3 minutes to about 4 minutes, about 3 minutes to about 5 minutes, about 3 minutes to about 6 minutes, about 3 minutes to about 7 minutes, about 3 minutes to about 7.5 minutes, about 3 minutes to about 10 minutes, about 3 minutes to about 15 minutes, about 3.5 minutes to about 4 minutes, about 3.5 minutes to about 5 minutes, about 3.5 minutes to about 6 minutes, about 3.5WSGR Docket No. 52459-726.601 minutes to about 7 minutes, about 3.5 minutes to about 7.5 minutes, about 3.5 minutes to about 10 minutes, about 3.5 minutes to about 15 minutes, about 4 minutes to about 5 minutes, about 4 minutes to about 6 minutes, about 4 minutes to about 7 minutes, about 4 minutes to about 7.5 minutes, about 4 minutes to about 10 minutes, about 4 minutes to about 15 minutes, about 5 minutes to about 6 minutes, about 5 minutes to about 7 minutes, about 5 minutes to about 7.5 minutes, about 5 minutes to about 10 minutes, about 5 minutes to about 15 minutes, about 6 minutes to about 7 minutes, about 6 minutes to about 7.5 minutes, about 6 minutes to about 10 minutes, about 6 minutes to about 15 minutes, about 7 minutes to about 7.5 minutes, about 7 minutes to about 10 minutes, about 7 minutes to about 15 minutes, about 7.5 minutes to about 10 minutes, about 7.5 minutes to about 15 minutes, or about 10 minutes to about 15 minutes in the lysis buffer.

[0247] In some embodiments, the lysis buffer can be stable for a period of time prior to addition of a sample. In some embodiments, the lysis buffer can be stable for at least about 30 seconds, at least about 1 minute, at least about 2 minutes, at least about 5 minutes, at least about 10 minutes, at least about 15 minutes, at least about 20 minutes, at least about 25 minutes, at least about 30 minutes, at least about 60 minutes, at least about 90 minutes, at least about 120 minutes, at least about 12 hours, at least about 1 day, at least about 3 days, at least about 5 days, at least about 10 days, at least about 15 days, at least about 1 month, or at least about 3 months. In some embodiments, the sample is incubated for at most about 3 months, at most about 1 month, at most about 15 days, at most about 10 days, at most about 3 days, at most about 1 day, at most about 12 hours, at most about 120 minutes, at most about 90 minutes, at most about 60 minutes, at most about 50 minutes, at most about 40 minutes, at most about 30 minutes, at most about 25 minutes, at most about 20 minutes, at most about 15 minutes, at most about 12 minutes, at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6.5 minutes, at most about 6 minutes, at most about 5.5 minutes, at most about 5 minutes, at most about 4.5 minutes, at most about 4 minutes, at most about 3.5 minutes, at most about 3 minutes, at most about 2.5 minutes, at most about 2 minutes, at most about 1.5 minutes, at most about 1 minute, at most about 30 seconds, or at most about 15 seconds.

[0248] In some embodiments, the lysis buffer can be stable for about 1 hour to about 48 hours. In some embodiments, the lysis buffer can be stable for about 1 hour to about 2 hours, about 1 hour to about 3 hours, about 1 hour to about 4 hours, about 1 hour to about 5 hours, about 1 hour to about 7 hours, about 1 hour to about 10 hours, about 1 hour to about 12 hours, about 1 hour to about 18 hours, about 1 hour to about 24 hours, about 1 hour to about 36 hours, about 1 hour toWSGR Docket No. 52459-726.601 about 48 hours, about 2 hours to about 3 hours, about 2 hours to about 4 hours, about 2 hours to about 5 hours, about 2 hours to about 7 hours, about 2 hours to about 10 hours, about 2 hours to about 12 hours, about 2 hours to about 18 hours, about 2 hours to about 24 hours, about 2 hours to about 36 hours, about 2 hours to about 48 hours, about 3 hours to about 4 hours, about 3 hours to about 5 hours, about 3 hours to about 7 hours, about 3 hours to about 10 hours, about 3 hours to about 12 hours, about 3 hours to about 18 hours, about 3 hours to about 24 hours, about 3 hours to about 36 hours, about 3 hours to about 48 hours, about 4 hours to about 5 hours, about 4 hours to about 7 hours, about 4 hours to about 10 hours, about 4 hours to about 12 hours, about 4 hours to about 18 hours, about 4 hours to about 24 hours, about 4 hours to about 36 hours, about4 hours to about 48 hours, about 5 hours to about 7 hours, about 5 hours to about 10 hours, about5 hours to about 12 hours, about 5 hours to about 18 hours, about 5 hours to about 24 hours, about 5 hours to about 36 hours, about 5 hours to about 48 hours, about 7 hours to about 10 hours, about 7 hours to about 12 hours, about 7 hours to about 18 hours, about 7 hours to about 24 hours, about 7 hours to about 36 hours, about 7 hours to about 48 hours, about 10 hours to about 12 hours, about 10 hours to about 18 hours, about 10 hours to about 24 hours, about 10 hours to about 36 hours, about 10 hours to about 48 hours, about 12 hours to about 18 hours, about 12 hours to about 24 hours, about 12 hours to about 36 hours, about 12 hours to about 48 hours, about 18 hours to about 24 hours, about 18 hours to about 36 hours, about 18 hours to about 48 hours, about 24 hours to about 36 hours, about 24 hours to about 48 hours, or about 36 hours to about 48 hours.

[0249] In some embodiments, the lysis buffer may be used as a storage buffer for transportation. In some embodiments, the sample may be stored in the lysis buffer for at least about 15 seconds, at least about 30 seconds, at least about 1 minute, at least about 1.5 minutes, at least about 2 minutes, at least about 2.5 minutes, at least about 3 minutes, at least about 3.5 minutes, at least about 4 minutes, at least about 4.5 minutes, at least about 5 minutes, at least about 5.5 minutes, at least about 6 minutes, at least about 6.5 minutes, at least about 7 minutes, at least about 8 minutes, at least about 9 minutes, at least about 10 minutes, at least about 12 minutes, at least about 15 minutes, at least about 20 minutes, at least about 25 minutes, at least about 30 minutes, at least about 40 minutes, at least about 50 minutes, at least about 60 minutes, at least about 90 minutes, or at least about 120 minutes. In some embodiments, the sample may be stored in the lysis buffer for at most about 120 minutes, at most about 90 minutes, at most about 60 minutes, at most about 50 minutes, at most about 40 minutes, at most about 30 minutes, at most about 25 minutes, at most about 20 minutes, at most about 15 minutes, at most about 12WSGR Docket No. 52459-726.601 minutes, at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6.5 minutes, at most about 6 minutes, at most about 5.5 minutes, at most about 5 minutes, at most about 4.5 minutes, at most about 4 minutes, at most about 3.5 minutes, at most about 3 minutes, at most about 2.5 minutes, at most about 2 minutes, at most about 1.5 minutes, at most about 1 minute, at most about 30 seconds, or at most about 15 seconds.

[0250] In some embodiments, the sample may be refrigerated prior to contacting the lysis buffer.

[0251] The sample may be heated after incubation in the lysis buffer. The sample may be heated before incubation in the lysis buffer. In some embodiments, the sample is heated at a constant temperature for a period of time. In some embodiments, the constant temperature is at least about 30°C, at least about 40°C, at least about 50°C, at least about 60°C, at least about 70°C, at least about 75°C, at least about 80°C, at least about 85°C, at least about 90°C, at least about 91°C, at least about 92°C, at least about 93°C, at least about 94°C, at least about 95°C, at least about 96°C, at least about 97°C, at least about 98°C, at least about 99°C, at least about 100°C, at least about 105°C, at least about 110°C, at least about 115°C, at least about 120°C, at least about 125°C, at least about 130°C, at least about 140°C, or at least about 150°C. In some embodiments, the constant temperature is at most about 150°C, at most about 140°C, at most about 130°C, at most about 125°C, at most about 120°C, at most about 115°C, at most about 100°C, at most about 105 °C, at most about 100°C, at most about 99°C, at most about 98°C, at most about 97°C, at most about 96°C, at most about 95°C, at most about 94°C, at most about 93°C, at most about 92°C, at most about 91°C, at most about 90°C, at most about 85°C, at most about 80°C, at most about 75°C, at most about 70°C, at most about 60°C, at most about 50°C, at most about 40°C, or at most about 30°C.

[0252] In some embodiments, the constant temperature is about 50°C to about 120°C. In some embodiments, the constant temperature is at most about 120°C. In some embodiments, the constant temperature is about 50°C to about 60°C, about 50°C to about 70°C, about 50°C to about 80°C, about 50°C to about 85°C, about 50°C to about 90°C, about 50°C to about 95°C, about 50°C to about 100°C, about 50°C to about 105°C, about 50°C to about 110°C, about 50°C to about 115°C, about 50°C to about 120°C, about 60°C to about 70°C, about 60°C to about 80°C, about 60°C to about 85°C, about 60°C to about 90°C, about 60°C to about 95°C, about 60°C to about 100°C, about 60°C to about 105°C, about 60°C to about 110°C, about 60°C to about 115°C, about 60°C to about 120°C, about 70°C to about 80°C, about 70°C to about 85°C,WSGR Docket No. 52459-726.601 about 70°C to about 90°C, about 70°C to about 95°C, about 70°C to about 100°C, about 70°C to about 105°C, about 70°C to about 110°C, about 70°C to about 115°C, about 70°C to about 120°C, about 80°C to about 85°C, about 80°C to about 90°C, about 80°C to about 95°C, about 80°C to about 100°C, about 80°C to about 105°C, about 80°C to about 110°C, about 80°C to about 115°C, about 80°C to about 120°C, about 85°C to about 90°C, about 85°C to about 95°C, about 85°C to about 100°C, about 85°C to about 105°C, about 85°C to about 110°C, about 85°C to about 115°C, about 85°C to about 120°C, about 90°C to about 95°C, about 90°C to about 100°C, about 90°C to about 105°C, about 90°C to about 110°C, about 90°C to about 115°C, about 90°C to about 120°C, about 95°C to about 100°C, about 95°C to about 105°C, about 95°C to about 110°C, about 95°C to about 115°C, about 95°C to about 120°C, about 100°C to about 105°C, about 100°C to about 110°C, about 100°C to about 115°C, about 100°C to about 120°C, about 105°C to about 110°C, about 105°C to about 115°C, about 105°C to about 120°C, about 110°C to about 115 °C, about 110°C to about 120°C, or about 115 °C to about 120°C.

[0253] In some embodiments, the sample is heated at a constant temperature for a period of time. In some embodiments, the sample is heated at a constant temperature for at least about 15 seconds, at least about 30 seconds, at least about 1 minute, at least about 1.5 minutes, at least about 2 minutes, at least about 2.5 minutes, at least about 3 minutes, at least about 3.5 minutes, at least about 4 minutes, at least about 4.5 minutes, at least about 5 minutes, at least about 5.5 minutes, at least about 6 minutes, at least about 6.5 minutes, at least about 7 minutes, at least about 8 minutes, at least about 9 minutes, at least about 10 minutes, at least about 12 minutes, or at least about 15 minutes. In some embodiments, the sample is incubated for at most about 15 minutes, at most about 12 minutes, at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6.5 minutes, at most about 6 minutes, at most about 5.5 minutes, at most about 5 minutes, at most about 4.5 minutes, at most about 4 minutes, at most about 3.5 minutes, at most about 3 minutes, at most about 2.5 minutes, at most about 2 minutes, at most about 1.5 minutes, at most about 1 minute, at most about 30 seconds, or at most about 15 seconds.

[0254] In some embodiments, the sample is heated at a constant temperature for about 1 minute to about 15 minutes. In some embodiments, the sample is heated at a constant temperature for about 1 minute to about 2 minutes, about 1 minute to about 2.5 minutes, about 1 minute to about 3 minutes, about 1 minute to about 3.5 minutes, about 1 minute to about 4 minutes, about 1 minute to about 5 minutes, about 1 minute to about 6 minutes, about 1 minute to about 7 minutes, about 1 minute to about 7.5 minutes, about 1 minute to about 10 minutes, about 1WSGR Docket No. 52459-726.601 minute to about 15 minutes, about 2 minutes to about 2.5 minutes, about 2 minutes to about 3 minutes, about 2 minutes to about 3.5 minutes, about 2 minutes to about 4 minutes, about 2 minutes to about 5 minutes, about 2 minutes to about 6 minutes, about 2 minutes to about 7 minutes, about 2 minutes to about 7.5 minutes, about 2 minutes to about 10 minutes, about 2 minutes to about 15 minutes, about 2.5 minutes to about 3 minutes, about 2.5 minutes to about3.5 minutes, about 2.5 minutes to about 4 minutes, about 2.5 minutes to about 5 minutes, about2.5 minutes to about 6 minutes, about 2.5 minutes to about 7 minutes, about 2.5 minutes to about7.5 minutes, about 2.5 minutes to about 10 minutes, about 2.5 minutes to about 15 minutes, about 3 minutes to about 3.5 minutes, about 3 minutes to about 4 minutes, about 3 minutes to about 5 minutes, about 3 minutes to about 6 minutes, about 3 minutes to about 7 minutes, about 3 minutes to about 7.5 minutes, about 3 minutes to about 10 minutes, about 3 minutes to about 15 minutes, about 3.5 minutes to about 4 minutes, about 3.5 minutes to about 5 minutes, about3.5 minutes to about 6 minutes, about 3.5 minutes to about 7 minutes, about 3.5 minutes to about7.5 minutes, about 3.5 minutes to about 10 minutes, about 3.5 minutes to about 15 minutes, about 4 minutes to about 5 minutes, about 4 minutes to about 6 minutes, about 4 minutes to about 7 minutes, about 4 minutes to about 7.5 minutes, about 4 minutes to about 10 minutes, about 4 minutes to about 15 minutes, about 5 minutes to about 6 minutes, about 5 minutes to about 7 minutes, about 5 minutes to about 7.5 minutes, about 5 minutes to about 10 minutes, about 5 minutes to about 15 minutes, about 6 minutes to about 7 minutes, about 6 minutes to about 7.5 minutes, about 6 minutes to about 10 minutes, about 6 minutes to about 15 minutes, about 7 minutes to about 7.5 minutes, about 7 minutes to about 10 minutes, about 7 minutes to about 15 minutes, about 7.5 minutes to about 10 minutes, about 7.5 minutes to about 15 minutes, or about 10 minutes to about 15 minutes.

[0255] In some embodiments, the sample is heated at a cyclic temperature for a period of time. In some embodiments, the cyclic temperature comprises a range of temperatures. In some embodiments, the cyclic temperature is about 30°C to about 120°C. In some embodiments, the cyclic temperature is about 30°C to about 40°C, about 30°C to about 50°C, about 30°C to about 60°C, about 30°C to about 70°C, about 30°C to about 80°C, about 30°C to about 85°C, about 30°C to about 90°C, about 30°C to about 95°C, about 30°C to about 100°C, about 30°C to about 110°C, about 30°C to about 120°C, about 40°C to about 50°C, about 40°C to about 60°C, about 40°C to about 70°C, about 40°C to about 80°C, about 40°C to about 85°C, about 40°C to about 90°C, about 40°C to about 95°C, about 40°C to about 100°C, about 40°C to about 110°C, about 40°C to about 120°C, about 50°C to about 60°C, about 50°C to about 70°C, about 50°C to aboutWSGR Docket No. 52459-726.60180°C, about 50°C to about 85°C, about 50°C to about 90°C, about 50°C to about 95°C, about 50°C to about 100°C, about 50°C to about 110°C, about 50°C to about 120°C, about 60°C to about 70°C, about 60°C to about 80°C, about 60°C to about 85°C, about 60°C to about 90°C, about 60°C to about 95°C, about 60°C to about 100°C, about 60°C to about 110°C, about 60°C to about 120°C, about 70°C to about 80°C, about 70°C to about 85°C, about 70°C to about 90°C, about 70°C to about 95°C, about 70°C to about 100°C, about 70°C to about 110°C, about 70°C to about 120°C, about 80°C to about 85°C, about 80°C to about 90°C, about 80°C to about 95°C, about 80°C to about 100°C, about 80°C to about 110°C, about 80°C to about 120°C, about 85°C to about 90°C, about 85°C to about 95°C, about 85°C to about 100°C, about 85°C to about 110°C, about 85°C to about 120°C, about 90°C to about 95°C, about 90°C to about 100°C, about 90°C to about 110°C, about 90°C to about 120°C, about 95°C to about 100°C, about 95°C to about 110°C, about 95°C to about 120°C, about 100°C to about 110°C, about 100°C to about 120°C, or about 110°C to about 120°C.

[0256] In some embodiments, the cyclic temperature is at most about 115°C. In some embodiments, the cyclic temperature is about 80°C to about 83°C, about 80°C to about 85°C, about 80°C to about 87°C, about 80°C to about 90°C, about 80°C to about 93 °C, about 80°C to about 95°C, about 80°C to about 97°C, about 80°C to about 100°C, about 80°C to about 105°C, about 80°C to about 110°C, about 80°C to about 115°C, about 83°C to about 85°C, about 83°C to about 87°C, about 83°C to about 90°C, about 83°C to about 93°C, about 83°C to about 95°C, about 83°C to about 97°C, about 83°C to about 100°C, about 83°C to about 105°C, about 83°C to about 110°C, about 83°C to about 115°C, about 85°C to about 87°C, about 85°C to about 90°C, about 85°C to about 93°C, about 85°C to about 95°C, about 85°C to about 97°C, about 85°C to about 100°C, about 85°C to about 105°C, about 85°C to about 110°C, about 85°C to about 115°C, about 87°C to about 90°C, about 87°C to about 93°C, about 87°C to about 95°C, about 87°C to about 97°C, about 87°C to about 100°C, about 87°C to about 105°C, about 87°C to about 110°C, about 87°C to about 115 °C, about 90°C to about 93 °C, about 90°C to about 95°C, about 90°C to about 97°C, about 90°C to about 100°C, about 90°C to about 105°C, about 90°C to about 110°C, about 90°C to about 115°C, about 93°C to about 95°C, about 93°C to about 97°C, about 93°C to about 100°C, about 93°C to about 105°C, about 93°C to about 110°C, about 93°C to about 115°C, about 95°C to about 97°C, about 95°C to about 100°C, about 95°C to about 105°C, about 95°C to about 110°C, about 95°C to about 115°C, about 97°C to about 100°C, about 97°C to about 105 °C, about 97° C to about 110°C, about 97°C to aboutWSGR Docket No. 52459-726.601115°C, about 100°C to about 105°C, about 100°C to about 110°C, about 100°C to about 115°C, about 105°C to about 110°C, about 105°C to about 115°C, or about 110°C to about 115°C.

[0257] In some embodiments, the sample can be sonicated. Sonication may agitate particles in the sample. Without wishing to be bound by theory, sonication may assist in cell lysis during the sample processing. In some embodiments, the sample is sonicated for at least about 15 seconds, at least about 30 seconds, at least about 1 minute, at least about 1.5 minutes, at least about 2 minutes, at least about 2.5 minutes, at least about 3 minutes, at least about 3.5 minutes, at least about 4 minutes, at least about 4.5 minutes, at least about 5 minutes, at least about 5.5 minutes, at least about 6 minutes, at least about 6.5 minutes, at least about 7 minutes, at least about 8 minutes, at least about 9 minutes, or at least about 10 minutes.

[0258] In some embodiments, sonicating the sample occurs prior to heating the sample. In some embodiments, sonicating the sample occurs subsequent to heating the sample. In some embodiments, sonicating the sample occurs concurrent to heating the sample. In some embodiments, sonicating occurs without heating the sample.

[0259] After the sample is incubated at room temperature and / or heated, the sample may be contacted with a recovery buffer. The recovery buffer may add to the enzyme stability and / or robustness. In some aspects, the sample is contacted with a recovery buffer comprising a solubilizer and a cyclodextrin.

[0260] In some embodiments, the solubilizer comprises a polysorbate. The polysorbate may be polyoxyethylene (20) sorbitan monooleate (e.g., polysorbate 80), polyoxyethylene (20) sorbitan monolaurate (e.g., polysorbate 20), polyoxyethylene (20) sorbitan monopalmitate (e.g., polysorbate 40), polyoxyethylene (20) sorbitan monostearate (e.g., polysorbate 60), or a functional variant thereof. In some embodiments, the solubilizer is a Tergitol™ surfactant, a Triton™ surfactant, or a Igepal® surfactant. In some embodiments, the solubilizer is an alkoxylate or a cocamide. In some embodiments, the solubilizer is decyl glucoside, alkyl polyglycoside, lauryl glucoside, sorbitan tristearate, or a niosome. The solubilizer may mix with the detergent of the present composition. In some embodiments, the solubilizer is capable of forming micelles comprising the detergent of the present application.

[0261] In some embodiments, the solubilizer is polysorbate 80. In some embodiments, the concentration (e.g., final concentration) of the solubilizer in the mixture in the presence of the sample is at least about 0.05% v / v, at least about 0.1% v / v, at least about 0.5% v / v, at least about 1% v / v, at least about 5% v / v, at least about 10% v / v, at least about 15% v / v, at least about 20% v / v, at least about 22.5% v / v, at least about 25% v / v, at least about 27.5% v / v, at least about 30%WSGR Docket No. 52459-726.601N / N, at least about 32.5% v / v, at least about 35% v / v, at least about 37.5% v / v, at least about 40% v / v, at least about 42.5% v / v, at least about 45% v / v, at least about 47.5% v / v, at least about 50% v / v, at least about 52.5% v / v, at least about 55% v / v, at least about 57.5% v / v, at least about 60% v / v, at least about 70% v / v, or at least about 75% v / v.

[0262] In some embodiments, the concentration (e.g., final concentration) of the solubilizer in the mixture in the presence of the sample is at most about 75% v / v, at most about 70% v / v, at most about 65% v / v, at most about 60% v / v, at most about 57.5% v / v, at most about 55% v / v, at most about 52.5% v / v, at most about 50% v / v, at most about 47.5% v / v, at most about 45% v / v, at most about 42.5% v / v, at most about 40% v / v, at most about 37.5% v / v, at most about 35% v / v, at most about 32.5% v / v, at most about 30% v / v, at most about 27.5% v / v, at most about 25% v / v, at most about 22.5% v / v, at most about 20% v / v, at most about 15% v / v, at most about 10% v / v, at most about 5% v / v, at most about 1% v / v, at most about 0.5% v / v, at most about 0.1% v / v, or at most about 0.05% v / v.

[0263] In some embodiments, the cyclodextrin comprises cyclodextrin comprises (2- hydroxypropyl) P-cyclodextrin, (2-hydroxypropyl) y-cyclodextrin, (2-hydroxypropyl)-a- cyclodextrin, 3 A-amino-3 A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-P- cyclodextrin, 6-O-alpha-D-Maltosyl-P-cyclodextrin, 2,6-Di-O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A- amino-3A-deoxy-(2AS,3 AS)-y-cyclodextrin hydrate, or any combination thereof.

[0264] In some embodiments, the cyclodextrin is present at a final concentration mixed with the sample that is effective for isolating the detergent within the composition of the present invention. In some embodiments, the concentration (e.g., final concentration) of the cyclodextrin in the mixture in the presence of the sample is at least about, at most about, or about 0.05 mM, 0.1 mM, 0.5 mM, 1.0 mM, 5.0 mM, 10.0 mM, 15.0 mM, 20.0 mM, 25.0 mM, 30.0 mM, 35.0 mM, 40.0 mM, 50.0 mM, 55.0 mM, 60.0 mM, 65.0 mM, 70.0 mM, 75.0 mM, 80.0 mM, 85.0 mM, 90.0 mM, 95.0 mM, 100.0 mM, 125.0 mM, 150.0 mM, 175.0 mM, 200.0 mM, 250.0 mM, 300.0 mM, or a range between any two of these values.

[0265] In some embodiments, the final volume of the recovery buffer is at least about, at most about, or about 100 pl, 200 pl, 300 pl, 400 pl, 450 pl, 500 pl, 550 pl, 600 pl, 650 pl, 700 pl, 750 pl, 800 pl, 900 pl, 1000 pl, or a range between any two of these values.

[0266] In some embodiments, the sample is mixed with the recovery buffer. In some embodiments, mixing of the sample and recovery buffer is by a vortex and / or by hand. In some embodiments, the mixing of the sample and the recovery buffer is performed by an automatedWSGR Docket No. 52459-726.601 instrument, a consumable, a microfluidic system, or a millifluidic system. In some embodiments, the mixing of the sample and the recovery buffer is performed until the sample and the recovery buffer are mixed to homogeneity.

[0267] In some embodiments, the lysis buffer may be frozen to stabilize the solution. In some embodiments, the lysis buffer may be frozen at a temperature of between about -50°C to 0°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -50°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -40°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -30°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -25°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -20°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -15°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -10°C. In some embodiments, the lysis buffer may be frozen at a temperature of about -5°C. In some embodiments, the lysis buffer may be frozen at a temperature of about 0°C.

[0268] After freezing the lysis buffer, the buffer may be thawed and mixed with the sample. In some embodiments, the efficiency of the thawed lysis buffer is tested and compared with the efficiency of an unfrozen lysis buffer. In some embodiments, the efficiency of the thawed lysis buffer is similar to the efficiency of the unfrozen lysis buffer.

[0269] In some embodiments, the recovery buffer may be frozen to stabilize the solution. In some embodiments, the recovery buffer may be frozen at a temperature of between about - 120°C to 0°C. In some embodiments, the recovery buffer may be frozen at a temperature of about -80°C. In some embodiments, the recovery buffer may be frozen at a temperature of about -120°C to about 0°C. In some embodiments, the recovery buffer may be frozen at a temperature of at least about -120°C. In some embodiments, the recovery buffer may be frozen at a temperature of at most about 0°C. In some embodiments, the recovery buffer may be frozen at a temperature of about -120°C to about -100°C, about -120°C to about -90°C, about -120°C to about -80°C, about -120°C to about -70°C, about -120°C to about -60°C, about -120°C to about -50°C, about -120°C to about -40°C, about -120°C to about -30°C, about -120°C to about -20°C, about -120°C to about -10°C, about -120°C to about 0°C, about -100°C to about -90°C, about - 100°C to about -80°C, about -100°C to about -70°C, about -100°C to about -60°C, about -100°C to about -50°C, about -100°C to about -40°C, about -100°C to about -30°C, about -100°C to about -20°C, about -100°C to about -10°C, about -100°C to about 0°C, about -90°C to about - 80°C, about -90°C to about -70°C, about -90°C to about -60°C, about -90°C to about -50°C,WSGR Docket No. 52459-726.601 about -90°C to about -40°C, about -90°C to about -30°C, about -90°C to about -20°C, about - 90°C to about -10°C, about -90°C to about 0°C, about -80°C to about -70°C, about -80°C to about -60°C, about -80°C to about -50°C, about -80°C to about -40°C, about -80°C to about - 30°C, about -80°C to about -20°C, about -80°C to about -10°C, about -80°C to about 0°C, about -70°C to about -60°C, about -70°C to about -50°C, about -70°C to about -40°C, about -70°C to about -30°C, about -70°C to about -20°C, about -70°C to about -10°C, about -70°C to about 0°C, about -60°C to about -50°C, about -60°C to about -40°C, about -60°C to about -30°C, about -60°C to about -20°C, about -60°C to about -10°C, about -60°C to about 0°C, about -50°C to about -40°C, about -50°C to about -30°C, about -50°C to about -20°C, about -50°C to about - 10°C, about -50°C to about 0°C, about -40°C to about -30°C, about -40°C to about -20°C, about -40°C to about -10°C, about -40°C to about 0°C, about -30°C to about -20°C, about -30°C to about -10°C, about -30°C to about 0°C, about -20°C to about -10°C, about -20°C to about 0°C, or about -10°C to about 0°C.

[0270] After freezing the recovery buffer, the buffer may be thawed and mixed with the sample. In some embodiments, the efficiency of the thawed recovery buffer is tested and compared with the efficiency of an unfrozen recovery buffer. In some embodiments, the efficiency of the thawed recovery buffer is similar to the efficiency of the unfrozen recovery buffer.

[0271] In some embodiments, the recovery buffer may be lyophilized.

[0272] After the sample has been contacted by the recovery buffer, the sample may be a processed sample. Without wishing to be bound by theory, the sample processing methods of the present disclosure may reduce the total preparation time of an unprocessed sample to a processed sample. In some embodiments, the sample processing of the present disclosure shortens the processing time of a sample by at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 70%, or at least about 80% compared to the processing time of a sample by a different sample processing method (e.g., a sample processing method with SDS, polysorbate 80, or a cyclodextrin used individually). A different sample processing methods can comprise alternative reagents, including, but not limited to, isopropanol and / or ethanol. In some embodiments, the sample processing of the present disclosure shortens the processing time of a sample by at most about 75%, at most about 70%, at most about 65%, at most about 60%, at most about 55%, at most about 50%, at most about 45%, at most about 40%, at most about 35%,WSGR Docket No. 52459-726.601 at most about 30%, at most about 25%, at most about 20%, at most about 15%, at most about 10%, at most about 5%, or at most about 3% compared to the processing time of a sample by a different sample processing method.

[0273] In some embodiments, the sample processing of the present disclosure shortens the processing time of a sample by about 2% to about 75%. In some embodiments, the sample processing of the present disclosure shortens the processing time of a sample by at most about 75%. In some embodiments, the sample processing of the present disclosure shortens the processing time of a sample by about 2% to about 3%, about 2% to about 5%, about 2% to about 7%, about 2% to about 10%, about 2% to about 15%, about 2% to about 20%, about 2% to about 30%, about 2% to about 40%, about 2% to about 50%, about 2% to about 60%, about 2% to about 75%, about 3% to about 5%, about 3% to about 7%, about 3% to about 10%, about 3% to about 15%, about 3% to about 20%, about 3% to about 30%, about 3% to about 40%, about 3% to about 50%, about 3% to about 60%, about 3% to about 75%, about 5% to about 7%, about 5% to about 10%, about 5% to about 15%, about 5% to about 20%, about 5% to about 30%, about 5% to about 40%, about 5% to about 50%, about 5% to about 60%, about 5% to about 75%, about 7% to about 10%, about 7% to about 15%, about 7% to about 20%, about 7% to about 30%, about 7% to about 40%, about 7% to about 50%, about 7% to about 60%, about 7% to about 75%, about 10% to about 15%, about 10% to about 20%, about 10% to about 30%, about 10% to about 40%, about 10% to about 50%, about 10% to about 60%, about 10% to about 75%, about 15% to about 20%, about 15% to about 30%, about 15% to about 40%, about 15% to about 50%, about 15% to about 60%, about 15% to about 75%, about 20% to about 30%, about 20% to about 40%, about 20% to about 50%, about 20% to about 60%, about 20% to about 75%, about 30% to about 40%, about 30% to about 50%, about 30% to about 60%, about 30% to about 75%, about 40% to about 50%, about 40% to about 60%, about 40% to about 75%, about 50% to about 60%, about 50% to about 75%, or about 60% to about 75%.

[0274] In an aspect, the present disclosure provides a method of processing a sample, the method comprising: (a) contacting the sample with a lysis buffer comprising a detergent; (b) incubating the sample at a first temperature or temperature range for a first time period; (c) heating the sample at a second temperature or temperature range for a second time period; and (d) contacting the sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing the sample to generate a processed sample in a mixture comprising the detergent, the solubilizer and the cyclodextrin.WSGR Docket No. 52459-726.601

[0275] In some embodiments, heating the sample in (c) further comprises heating the sample to the second temperature, cooling down the sample, and heating the sample to the second temperature after cooling down. In some embodiments, the sample may be cooled down to room temperature. In some embodiments, the sample may be cooled down to a temperature below room temperature.

[0276] In some embodiments, the sample in (b) may be incubated at a temperature of at least about -10°C, at least about -5°C, at least about 0°C, at least about 5°C, at least about 10°C, at least about 12°C, at least about 14°C, at least about 16°C, at least about 18°C, at least about 20°C, at least about 22°C, at least about 24°C, at least about 26°C, at least about 28°C, at least about 30°C, at least about 35°C, at least about 40°C, at least about 45°C, or at least about 50°C.

[0277] In some embodiments, the sample in (b) may be incubated at a temperature of at most about 50°C, at most about 45°C, at most about 40°C, at most about 35°C, at most about 30°C, at most about 28°C, at most about 26°C, at most about 24°C, at most about 22°C, at most about 20°C, at most about 18°C, at most about 16°C, at most about 14°C, at most about 12°C, at most about 10°C, at most about 5°C, at most about 0°C, at most about -5°C, or at most about -10°C.

[0278] In some embodiments, the sample in (b) may be incubated at a temperature of at about - 10°C to about 50°C. In some embodiments, the sample in (b) may be incubated at a temperature at about -10°C to about 0°C, about -10°C to about 10°C, about -10°C to about 12°C, about - 10°C to about 15 °C, about -10°C to about 17°C, about -10°C to about 20°C, about -10°C to about 22°C, about -10°C to about 25°C, about -10°C to about 27°C, about -10°C to about 30°C, about -10°C to about 50°C, about 0°C to about 10°C, about 0°C to about 12°C, about 0°C to about 15°C, about 0°C to about 17°C, about 0°C to about 20°C, about 0°C to about 22°C, about 0°C to about 25°C, about 0°C to about 27°C, about 0°C to about 30°C, about 0°C to about 50°C, about 10°C to about 12°C, about 10°C to about 15°C, about 10°C to about 17°C, about 10°C to about 20°C, about 10°C to about 22°C, about 10°C to about 25°C, about 10°C to about 27°C, about 10°C to about 30°C, about 10°C to about 50°C, about 12°C to about 15°C, about 12°C to about 17°C, about 12°C to about 20°C, about 12°C to about 22°C, about 12°C to about 25°C, about 12°C to about 27°C, about 12°C to about 30°C, about 12°C to about 50°C, about 15°C to about 17°C, about 15°C to about 20°C, about 15°C to about 22°C, about 15°C to about 25°C, about 15°C to about 27°C, about 15°C to about 30°C, about 15°C to about 50°C, about 17°C to about 20°C, about 17°C to about 22°C, about 17°C to about 25°C, about 17°C to about 27°C, about 17°C to about 30°C, about 17°C to about 50°C, about 20°C to about 22°C, about 20°C to about 25°C, about 20°C to about 27°C, about 20°C to about 30°C, about 20°C to about 50°C,WSGR Docket No. 52459-726.601 about 22°C to about 25°C, about 22°C to about 27°C, about 22°C to about 30°C, about 22°C to about 50°C, about 25°C to about 27°C, about 25°C to about 30°C, about 25°C to about 50°C, about 27°C to about 30°C, about 27°C to about 50°C, or about 30°C to about 50°C.

[0279] In some embodiments, the first time period is at least about 30 seconds, at least about 1 minute, at least about 2 minutes, at least about 3 minutes, at least about 4 minutes, at least about 5 minutes, at least about 10 minutes, at least about 15 minutes, at least about 20 minutes, at least about 25 minutes, at least about 30 minutes, at least about 45 minutes, at least about 60 minutes, at least about 2 hours, at least about 3 hours, at least about 4 hours, at least about 5 hours, at least about 10 hours, at least about 12 hours, at least about 24 hours, at least about 36 hours, at least about 48 hours, at least about 3 days, at least about 4 days, or at least about 5 days.

[0280] In some embodiments, the first time period is at most about 10 days, at most about 5 days, at most about 4 days, at most about 3 days, at most about 48 hours, at most about 36 hours, at most about 24 hours, at most about 12 hours, at most about 10 hours, at most about 5 hours, at most about 4 hours, at most about 3 hours, at most about 2 hours, at most about 60 minutes, at most about 45 minutes, at most about 30 minutes, at most about 25 minutes, at most about 20 minutes, at most about 15 minutes, at most about 10 minutes, at most about 5 minutes, at most about 4 minutes, at most about 3 minutes, at most about 2 minutes, at most about 1 minute, or at most about 30 seconds.

[0281] In some embodiments, a method of the present disclosure may comprise bead beating the sample. Bead beating can assist in cell lysis to agitate a sample with grinding media or beads. In some embodiments, a lysing matrix may be used for bead beating. In some embodiments, the lysing matrix may comprise silica, glass, ceramic, silicon carbide, zirconium silicate, garnet, stainless steel, and / or zirconium oxide.

[0282] In some embodiments, the second temperature of the method of the present disclosure is at least about, at most about, or about 30°C, 40°C, 50°C, 60°C, 70°C, 75°C, 80°C, 85°C, 90°C, 91°C, 92°C, 93°C, 94°C, 95°C, 96°C, 97°C, 98°C, 99°C, 100°C, 105°C, 110°C, 115°C, 120°C, 125°C, 130°C, 140°C, at least about 150°C, or a range between any two of these values.

[0283] In some embodiments, the second time period is at least about 30 seconds, at least about 1 minute, at least about 2 minutes, at least about 3 minutes, at least about 4 minutes, at least about 5 minutes, at least about 10 minutes, at least about 15 minutes, at least about 20 minutes, at least about 25 minutes, at least about 30 minutes, at least about 45 minutes, at least about 60 minutes, at least about 2 hours, at least about 3 hours, at least about 4 hours, or at least about 5 hours.WSGR Docket No. 52459-726.601

[0284] In some embodiments, the second time period is at most about 10 hours, at most about 5 hours, at most about 4 hours, at most about 3 hours, at most about 2 hours, at most about 60 minutes, at most about 45 minutes, at most about 30 minutes, at most about 25 minutes, at most about 20 minutes, at most about 15 minutes, at most about 10 minutes, at most about 5 minutes, at most about 4 minutes, at most about 3 minutes, at most about 2 minutes, at most about 1 minute, or at most about 30 seconds.

[0285] Subsequent to the recovery buffer, the processed sample may be contacted with a reaction mixture. In some embodiments, the reaction mixture comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, and / or a probe. In some embodiments,

[0286] In some embodiments, the thermostable enzyme comprises a large fragment of a Bacillus stearothermophilus polymerase, an exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and / or any mutants thereof. In some embodiments, the thermostable enzyme comprises a DNA polymerase. In some embodiments, the thermostable enzymes comprises a Taq DNA polymerase. In some embodiments, the thermostable enzyme comprises a DNA-dependent DNA polymerase. In some embodiments, the thermostable enzyme comprises a strand-displacing DNA polymerase. In some embodiments, the reaction mixture stabilizes enzymatic activity of the thermostable enzyme. A large fragment of a Bacillus stearothermophilus polymerase is the portion of the Bacillus stearothermophilusDNA polymerase that contains the 5' — » 3' polymerase activity, but lacks the 5' — >3' exonuclease domain. In some embodiments, the composition is configured to stabilize enzymatic activity of the thermostable enzyme for use during a nucleic acid amplification.

[0287] In some embodiments, the dNTPs of the reaction mixture comprise dATP, dCTP, dGTP, dTTP, and / or dUTP. In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is at least about 25 pM, at least about 50 pM, at least about 75 pM, at least about 100 pM, at least about 150 pM, at least about 200 pM, at least about 250 pM, at least about 300 pM, at least about 350 pM, at least about 400 pM, at least about 450 pM, at least about 500 pM, at least about 750 pM, at least about 1000 pM, at least about 1500 pM, at least about 2000 pM, at least about 2500 pM, at least about 3000 pM, at least about 3500 pM, at least about 4000 pM, at least about 4500 pM, at least about 5000 pM, at least about 6000 pM, at least about 7000 pM, at least about 8000 pM, at least about 9000 pM, or at least about 10000 pM.WSGR Docket No. 52459-726.601

[0288] In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is at most about 10000 pM, at most about 9000 pM, at most about 8000 pM, at most about 7000 pM, at most about 6000 pM, at most about 5000 pM, at most about 4500 pM, at most about 4000 pM, at most about 3500 pM, at most about 3000 pM, at most about 2500 pM, at most about 2000 pM, at most about 1500 pM, at most about 1000 pM, at most about 750 pM, at most about 500 pM, at most about 450 pM, at most about 400 pM, at most about 350 pM, at most about 300 pM, at most about 250 pM, at most about 200 pM, at most about 150 pM, at most about 100 pM, at most about 75 pM, at most about 50 pM, or at most about 25 pM.

[0289] In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is about 50 pM to about 7,500 pM. In some embodiments, a concentration of dNTPs in the reaction mixture when mixed with the sample is about 50 pM to about 100 pM, about 50 pM to about 250 pM, about 50 pM to about 500 pM, about 50 pM to about 750 pM, about 50 pM to about 1,000 pM, about 50 pM to about 1,250 pM, about 50 pM to about 1,500 pM, about 50 pM to about 2,000 pM, about 50 pM to about 4,000 pM, about 50 pM to about 5,000 pM, about 50 pM to about 7,500 pM, about 100 pM to about 250 pM, about 100 pM to about 500 pM, about 100 pM to about 750 pM, about 100 pM to about 1,000 pM, about 100 pM to about 1,250 pM, about 100 pM to about 1,500 pM, about 100 pM to about 2,000 pM, about 100 pM to about 4,000 pM, about 100 pM to about 5,000 pM, about 100 pM to about 7,500 pM, about 250 pM to about 500 pM, about 250 pM to about 750 pM, about 250 pM to about 1,000 pM, about 250 pM to about 1,250 pM, about 250 pM to about 1,500 pM, about 250 pM to about 2,000 pM, about 250 pM to about 4,000 pM, about 250 pM to about 5,000 pM, about 250 pM to about 7,500 pM, about 500 pM to about 750 pM, about 500 pM to about 1,000 pM, about 500 pM to about 1,250 pM, about 500 pM to about 1,500 pM, about 500 pM to about 2,000 pM, about 500 pM to about 4,000 pM, about 500 pM to about 5,000 pM, about 500 pM to about 7,500 pM, about 750 pM to about 1,000 pM, about 750 pM to about 1,250 pM, about 750 pM to about 1,500 pM, about 750 pM to about 2,000 pM, about 750 pM to about 4,000 pM, about 750 pM to about 5,000 pM, about 750 pM to about 7,500 pM, about 1,000 pM to about 1,250 pM, about 1,000 pM to about 1,500 pM, about 1,000 pM to about 2,000 pM, about 1,000 pM to about 4,000 pM, about 1,000 pM to about 5,000 pM, about 1,000 pM to about 7,500 pM, about 1,250 pM to about 1,500 pM, about 1,250 pM to about 2,000 pM, about 1,250 pM to about 4,000 pM, about 1,250 pM to about 5,000 pM, about 1,250 pM to about 7,500 pM, about 1,500 pM to about 2,000 pM, about 1,500 pM to about 4,000 pM, about 1,500 pM to about 5,000 pM, about 1,500 pM to about 7,500 pM, about 2,000 pM to about 4,000 pM,WSGR Docket No. 52459-726.601 about 2,000 pM to about 5,000 pM, about 2,000 pM to about 7,500 pM, about 4,000 pM to about 5,000 pM, about 4,000 pM to about 7,500 pM, or about 5,000 pM to about 7,500 pM.

[0290] In some embodiments, the primer or probe can be a stretch of nucleotides that hybridizes with a target nucleic acid sequence. In some embodiments, the primer is at least about 3 nucleotides, at least about 5 nucleotides, at least about 10 nucleotides, at least about 15 nucleotides, at least about 20 nucleotides, at least about 25 nucleotides, at least about 30 nucleotides, at least about 35 nucleotides, at least about 40 nucleotides, at least about 45 nucleotides, at least about 50 nucleotides, at least about 60 nucleotides, at least about 70 nucleotides, at least about 80 nucleotides, at least about 90 nucleotides, at least about 100 nucleotides, at least about 150 nucleotides, or at least about 200 nucleotides in length. In some embodiments, the primer is at most about 200 nucleotides, at most about 150 nucleotides, at most about 100 nucleotides, at most about 90 nucleotides, at most about 80 nucleotides, at most about 70 nucleotides, at most about 60 nucleotides, at most about 50 nucleotides, at most about 45 nucleotides, at most about 40 nucleotides, at most about 35 nucleotides, at most about 30 nucleotides, at most about 25 nucleotides, at most about 20 nucleotides, at most about 15 nucleotides, at most about 10 nucleotides, at most about 5 nucleotides, or at most about 3 nucleotides in length.

[0291] In some embodiments, the primer is about 3 nucleotides to about 100 nucleotides in length. In some embodiments, the primer is at most about 100 nucleotides. In some embodiments, the primer is about 3 nucleotides to about 5 nucleotides, about 3 nucleotides to about 10 nucleotides, about 3 nucleotides to about 20 nucleotides, about 3 nucleotides to about 30 nucleotides, about 3 nucleotides to about 40 nucleotides, about 3 nucleotides to about 50 nucleotides, about 3 nucleotides to about 60 nucleotides, about 3 nucleotides to about 70 nucleotides, about 3 nucleotides to about 80 nucleotides, about 3 nucleotides to about 90 nucleotides, about 3 nucleotides to about 100 nucleotides, about 5 nucleotides to about 10 nucleotides, about 5 nucleotides to about 20 nucleotides, about 5 nucleotides to about 30 nucleotides, about 5 nucleotides to about 40 nucleotides, about 5 nucleotides to about 50 nucleotides, about 5 nucleotides to about 60 nucleotides, about 5 nucleotides to about 70 nucleotides, about 5 nucleotides to about 80 nucleotides, about 5 nucleotides to about 90 nucleotides, about 5 nucleotides to about 100 nucleotides, about 10 nucleotides to about 20 nucleotides, about 10 nucleotides to about 30 nucleotides, about 10 nucleotides to about 40 nucleotides, about 10 nucleotides to about 50 nucleotides, about 10 nucleotides to about 60 nucleotides, about 10 nucleotides to about 70 nucleotides, about 10 nucleotides to about 80WSGR Docket No. 52459-726.601 nucleotides, about 10 nucleotides to about 90 nucleotides, about 10 nucleotides to about 100 nucleotides, about 20 nucleotides to about 30 nucleotides, about 20 nucleotides to about 40 nucleotides, about 20 nucleotides to about 50 nucleotides, about 20 nucleotides to about 60 nucleotides, about 20 nucleotides to about 70 nucleotides, about 20 nucleotides to about 80 nucleotides, about 20 nucleotides to about 90 nucleotides, about 20 nucleotides to about 100 nucleotides, about 30 nucleotides to about 40 nucleotides, about 30 nucleotides to about 50 nucleotides, about 30 nucleotides to about 60 nucleotides, about 30 nucleotides to about 70 nucleotides, about 30 nucleotides to about 80 nucleotides, about 30 nucleotides to about 90 nucleotides, about 30 nucleotides to about 100 nucleotides, about 40 nucleotides to about 50 nucleotides, about 40 nucleotides to about 60 nucleotides, about 40 nucleotides to about 70 nucleotides, about 40 nucleotides to about 80 nucleotides, about 40 nucleotides to about 90 nucleotides, about 40 nucleotides to about 100 nucleotides, about 50 nucleotides to about 60 nucleotides, about 50 nucleotides to about 70 nucleotides, about 50 nucleotides to about 80 nucleotides, about 50 nucleotides to about 90 nucleotides, about 50 nucleotides to about 100 nucleotides, about 60 nucleotides to about 70 nucleotides, about 60 nucleotides to about 80 nucleotides, about 60 nucleotides to about 90 nucleotides, about 60 nucleotides to about 100 nucleotides, about 70 nucleotides to about 80 nucleotides, about 70 nucleotides to about 90 nucleotides, about 70 nucleotides to about 100 nucleotides, about 80 nucleotides to about 90 nucleotides, about 80 nucleotides to about 100 nucleotides, or about 90 nucleotides to about 100 nucleotides in length.

[0292] In some embodiments, the primer is tagged with biotin or 6-carboxyfluorescein (FAM) for visualization on a lateral flow immunoassay strip.

[0293] In some embodiments, the reaction mixture is lyophilized.

[0294] In some embodiments, the methods and compositions of the present disclosure can provide for a faster time from obtaining a sample to generating a processed sample. In some embodiments, a time from obtaining the sample to generating the processed sample is at least about 30 seconds, at least about 1 minute, at least about 2 minutes, at least about 3 minutes, at least about 4 minutes, at least about 5 minutes, at least about 10 minutes, at least about 15 minutes, at least about 20 minutes, at least about 30 minutes, at least about 45 minutes, or at least about 1 hour. In some embodiments, a time from obtaining the sample to generating the processed sample is at most about 1 hour, at most about 45 minutes, at most about 30 minutes, at most about 20 minutes, at most about 15 minutes, at most about 10 minutes, at most about 5WSGR Docket No. 52459-726.601 minutes, at most about 4 minutes, at most about 3 minutes, at most about 2 minutes, at most about 1 minute, or at most about 30 seconds.

[0295] In some embodiments, a time from obtaining the sample to generating the processed sample is about 1 minute to about 45 minutes. In some embodiments, a time from obtaining the sample to generating the processed sample is at most about 45 minutes. In some embodiments, a time from obtaining the sample to generating the processed sample is about 1 minute to about 2 minutes, about 1 minute to about 3 minutes, about 1 minute to about 4 minutes, about 1 minute to about 5 minutes, about 1 minute to about 6 minutes, about 1 minute to about 7 minutes, about 1 minute to about 8 minutes, about 1 minute to about 10 minutes, about 1 minute to about 15 minutes, about 1 minute to about 30 minutes, about 1 minute to about 45 minutes, about 2 minutes to about 3 minutes, about 2 minutes to about 4 minutes, about 2 minutes to about 5 minutes, about 2 minutes to about 6 minutes, about 2 minutes to about 7 minutes, about 2 minutes to about 8 minutes, about 2 minutes to about 10 minutes, about 2 minutes to about 15 minutes, about 2 minutes to about 30 minutes, about 2 minutes to about 45 minutes, about 3 minutes to about 4 minutes, about 3 minutes to about 5 minutes, about 3 minutes to about 6 minutes, about 3 minutes to about 7 minutes, about 3 minutes to about 8 minutes, about 3 minutes to about 10 minutes, about 3 minutes to about 15 minutes, about 3 minutes to about 30 minutes, about 3 minutes to about 45 minutes, about 4 minutes to about 5 minutes, about 4 minutes to about 6 minutes, about 4 minutes to about 7 minutes, about 4 minutes to about 8 minutes, about 4 minutes to about 10 minutes, about 4 minutes to about 15 minutes, about 4 minutes to about 30 minutes, about 4 minutes to about 45 minutes, about 5 minutes to about 6 minutes, about 5 minutes to about 7 minutes, about 5 minutes to about 8 minutes, about 5 minutes to about 10 minutes, about 5 minutes to about 15 minutes, about 5 minutes to about 30 minutes, about 5 minutes to about 45 minutes, about 6 minutes to about 7 minutes, about 6 minutes to about 8 minutes, about 6 minutes to about 10 minutes, about 6 minutes to about 15 minutes, about 6 minutes to about 30 minutes, about 6 minutes to about 45 minutes, about 7 minutes to about 8 minutes, about 7 minutes to about 10 minutes, about 7 minutes to about 15 minutes, about 7 minutes to about 30 minutes, about 7 minutes to about 45 minutes, about 8 minutes to about 10 minutes, about 8 minutes to about 15 minutes, about 8 minutes to about 30 minutes, about 8 minutes to about 45 minutes, about 10 minutes to about 15 minutes, about 10 minutes to about 30 minutes, about 10 minutes to about 45 minutes, about 15 minutes to about 30 minutes, about 15 minutes to about 45 minutes, or about 30 minutes to about 45 minutes.WSGR Docket No. 52459-726.601

[0296] Following contact with the reaction mixture, the processed sample may then be subjected to a nucleic acid amplification method. Various nucleic acid amplification methods can be used with the compositions and methods disclosed herein to amplify target sequences in nucleic acid molecules. Methods for nucleic acid amplification include, but are not limited to, polymerase chain reaction (PCR), nucleic acid sequence based amplification (NASBA), oligonucleotide ligation assay (OLA), transcription mediated amplification (TMA), oligonucleotide extension and ligation, rolling circle amplification (RCA), and / or strand displacement amplification (SDA). The target sequence processed by the methods provided herein can be used for further downstream applications, e.g., isothermal amplifications. Exemplary isothermal amplification methods that can be used with the compositions and methods provided herein include, but are not limited to, helicase-dependent amplification (HD A), isothermal multiple displacement amplification (IMDA), loop-mediated isothermal amplification (LAMP), recombinase polymerase amplification (RPA), single primer isothermal amplification (SPIA), or strand displacement amplification (SDA).

[0297] In some embodiments, the nucleic acid amplification of the present disclosure comprises PCR or isothermal amplification. In some embodiments, a temperature is changed over the course of the nucleic acid amplification method. In some embodiments, the nucleic acid amplification comprises thermocycling the processed sample. In some embodiments, the nucleic acid amplification comprises keeping the processed sample at a constant temperature during the amplification.

[0298] In some embodiments, the reaction mixture includes probes to visualize amplified nucleic acid products. In some embodiments, the probes comprise strand displacement probes, intercalating fluorophores, pH-sensitive dyes, and / or detecting pyrophosphate products.

[0299] In some cases, the methods and compositions for processing nucleic acid molecule samples disclosed herein generate a higher yield of amplified nucleic acid products than a yield of amplified nucleic acid products generated by different sample processing methods or compositions. In some embodiments, the sample processing methods and compositions generate a yield of amplified nucleic acid products that is at least about 5 times, at least 10 times, at least about 50 times, at least 100 times, at least 150 times, at least 200 times, at least 250 times, at least 300 times, at least 350 times, at least 400 times, at least 450 times, at least 500 times, at least 600 times, at least 700 times, at least 800 times, at least 900 times, at least 1,000 times, at least 1,500 times, at least 2,000 times, at least 10 times, or at least 10,000 times greater than aWSGR Docket No. 52459-726.601 yield of amplified nucleic acid products from an otherwise identical sample processed by SDS, polysorbate 80, and / or a cyclodextrin individually.

[0300] In some embodiments, the sample processing methods and compositions generate a yield of amplified nucleic acid products that is about 3 times to about 1,000 times greater than a yield of amplified nucleic acid products from an otherwise identical sample processed by SDS, polysorbate 80, and / or a cyclodextrin individually. In some embodiments, the sample processing methods and compositions generate a yield of amplified nucleic acid products that is at most about 1,000 times. In some embodiments, the sample processing methods and compositions generate a yield of amplified nucleic acid products that is about 3 times to about 5 times, about 3 times to about 10 times, about 3 times to about 25 times, about 3 times to about 50 times, about 3 times to about 100 times, about 3 times to about 150 times, about 3 times to about 200 times, about 3 times to about 250 times, about 3 times to about 500 times, about 3 times to about 750 times, about 3 times to about 1,000 times, about 5 times to about 10 times, about 5 times to about 25 times, about 5 times to about 50 times, about 5 times to about 100 times, about 5 times to about 150 times, about 5 times to about 200 times, about 5 times to about 250 times, about 5 times to about 500 times, about 5 times to about 750 times, about 5 times to about 1,000 times, about 10 times to about 25 times, about 10 times to about 50 times, about 10 times to about 100 times, about 10 times to about 150 times, about 10 times to about 200 times, about 10 times to about 250 times, about 10 times to about 500 times, about 10 times to about 750 times, about 10 times to about 1,000 times, about 25 times to about 50 times, about 25 times to about 100 times, about 25 times to about 150 times, about 25 times to about 200 times, about 25 times to about 250 times, about 25 times to about 500 times, about 25 times to about 750 times, about 25 times to about 1,000 times, about 50 times to about 100 times, about 50 times to about 150 times, about 50 times to about 200 times, about 50 times to about 250 times, about 50 times to about 500 times, about 50 times to about 750 times, about 50 times to about 1,000 times, about 100 times to about 150 times, about 100 times to about 200 times, about 100 times to about 250 times, about 100 times to about 500 times, about 100 times to about 750 times, about 100 times to about 1,000 times, about 150 times to about 200 times, about 150 times to about 250 times, about 150 times to about 500 times, about 150 times to about 750 times, about 150 times to about 1,000 times, about 200 times to about 250 times, about 200 times to about 500 times, about 200 times to about 750 times, about 200 times to about 1,000 times, about 250 times to about 500 times, about 250 times to about 750 times, about 250 times to about 1,000 times, about 500 times to about 750 times, about 500 times to about 1,000 times, or about 750 times toWSGR Docket No. 52459-726.601 about 1,000 times greater than a yield of amplified nucleic acid products from an otherwise identical sample processed by SDS, polysorbate 80, and / or a cyclodextrin individually.

[0301] There may be a total time to produce a processed sample using the composition and / or methods described herein. In some embodiments, a total time to perform steps of processing a sample described herein (e.g., (a) contacting the sample with a lysis buffer comprising a detergent, (b) contacting the sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing the sample to generate a processed sample in a mixture comprising the detergent, the solubilizer, and the cyclodextrin) may be at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6 minutes, at most about 5 minutes, at most about 4 minutes, at most about 3 minutes, at most about 2 minutes, at most about 1 minute, at most about 50 seconds, at most about 40 seconds, at most about 30 seconds, at most about 20 seconds, at most about 15 seconds, at most about 10 seconds, or less than about 10 seconds.

[0302] In some embodiments, a time for processing a sample can comprise a time period from contacting the sample with a reaction mixture to generating a processed sample. The time period from contacting the sample with a reaction mixture to generating a processed sample may be at most about 10 minutes, at most about 9 minutes, at most about 8 minutes, at most about 7 minutes, at most about 6 minutes, at most about 5 minutes, at most about 4 minutes, at most about 3 minutes, at most about 2 minutes, at most about 1 minute, at most about 50 seconds, at most about 40 seconds, at most about 30 seconds, at most about 20 seconds, at most about 15 seconds, at most about 10 seconds, or less than about 10 seconds.

[0303] In some cases, the methods described herein may not comprise heating a sample. In some cases, the methods described herein may comprise heating a sample. A sample may be heated from about 50°C to about 120°C. In some embodiments, a sample may be heated from about 50°C to about 60°C, about 50°C to about 70°C, about 50°C to about 80°C, about 50°C to about 85°C, about 50°C to about 90°C, about 50°C to about 95°C, about 50°C to about 100°C, about 50°C to about 105°C, about 50°C to about 110°C, about 50°C to about 115°C, about 50°C to about 120°C, about 60°C to about 70°C, about 60°C to about 80°C, about 60°C to about 85°C, about 60°C to about 90°C, about 60°C to about 95°C, about 60°C to about 100°C, about 60°C to about 105°C, about 60°C to about 110°C, about 60°C to about 115°C, about 60°C to about 120°C, about 70°C to about 80°C, about 70°C to about 85°C, about 70°C to about 90°C, about 70°C to about 95°C, about 70°C to about 100°C, about 70°C to about 105°C, about 70°C to about 110°C, about 70°C to about 115°C, about 70°C to about 120°C, about 80°C to aboutWSGR Docket No. 52459-726.60185°C, about 80°C to about 90°C, about 80°C to about 95°C, about 80°C to about 100°C, about 80°C to about 105°C, about 80°C to about 110°C, about 80°C to about 115°C, about 80°C to about 120°C, about 85°C to about 90°C, about 85°C to about 95°C, about 85°C to about 100°C, about 85°C to about 105°C, about 85°C to about 110°C, about 85°C to about 115°C, about 85°C to about 120°C, about 90°C to about 95 °C, about 90°C to about 100°C, about 90°C to about 105°C, about 90°C to about 110°C, about 90°C to about 115°C, about 90°C to about 120°C, about 95°C to about 100°C, about 95°C to about 105°C, about 95°C to about 110°C, about 95°C to about 115°C, about 95°C to about 120°C, about 100°C to about 105°C, about 100°C to about 110°C, about 100°C to about 115°C, about 100°C to about 120°C, about 105°C to about 110°C, about 105°C to about 115°C, about 105°C to about 120°C, about 110°C to about 115°C, about 110°C to about 120°C, or about 115°C to about 120°C.

[0304] In some aspects, provided herein are methods of amplifying a sample. The method can comprise contacting a sample with a composition. The composition may be a sample processing buffer described herein. Contacting the sample with the sample processing buffer can generate a processed sample. The method can comprise contacting the processed sample with a composition (e.g., a sample amplification buffer). Contacting the processed sample with the sample amplification buffer can provide a condition for an amplification (e.g., a nucleic acid amplification) for the sample. The method can comprise subjected the processed sample to an amplification (e.g., a nucleic acid ampli...

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A composition for sample processing comprising: a detergent, a solubilizer, and a cyclodextrin, wherein said composition is configured to stabilize an enzyme during a nucleic acid amplification, and wherein said composition is configured to reduce and / or eliminate activity of a degrading nuclease.

2. The composition of claim 1, wherein said composition is configured to stabilize nucleic acids during said nucleic acid amplification.

3. The composition of claim 1 or 2, wherein said enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof.

4. The composition of any one of claims 1-3, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

5. The composition of any one of claims 1-4, wherein said solubilizer is a non-ionic surfactant.

6. The composition of any one of claims 1-4, wherein said solubilizer is a polysorbate, octylphenoxypoly ethoxy ethanol, 2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol, or a secondary alcohol ethoxylate.

7. The composition of claim 5 or 6, wherein said solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

8. The composition of any one of claims 1-5, wherein said detergent is part of a lysis buffer.

9. The composition of any one of claims 1-8, wherein said solubilizer and said cyclodextrin are part of a recovery buffer.

10. The composition of claim 9, wherein said lysis buffer and said recovery buffer are in a same mixture.

11. A composition for sample processing comprising a buffer comprising: a detergent, a solubilizer, and a cyclodextrin, wherein said composition is configured to stabilize an enzyme during a nucleic acid amplification, andwherein said composition is configured to inactivate a degrading nuclease.

12. The composition of claim 11, wherein said composition is configured to stabilize nucleic acids during said nucleic acid amplification.

13. The composition of claim 11 or 12, wherein said enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof.

14. The composition of any one of claims 11-13, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

15. The composition of any one of claims 11-14, wherein said solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

16. The composition of any one of claims 1-15, wherein said solubilizer and said cyclodextrin are configured to shorten a cycle threshold value or a time to result value in said nucleic acid amplification compared to a cycle threshold value or a time to result value in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually.

17. The composition of claim 16, wherein said cycle threshold value is at most 40 or said time to result value is at most 15 minutes.

18. The composition of any one of claims 1-17, wherein said solubilizer and said cyclodextrin are configured to decrease a coefficient of variation.

19. The composition of any one of claims 1-18, wherein said solubilizer and said cyclodextrin are configured to lower a limit of detection.

20. The composition of any one of claims 1-19, wherein said degrading nuclease is a ribonuclease.

21. The composition of any one of claims 8-10, wherein said lysis buffer has a pH value of 2 to 9.

22. The composition of any one of claims 8-10 and 21, wherein said lysis buffer further comprises a chelating agent.

23. The composition of claim 22, wherein said chelating agent is deferiprone, ethylenediamine, 1,10-Phenanthroline, oxalic acid, pentetic acid, deferasirox, deferoxamine, deferoxamine mesylate, or N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2- ethanediamine (TPEN).

24. The composition of any one of claims 8-10, 21 and 22, wherein said lysis buffer further comprises a reducing agent.

25. The composition of claim 24, wherein said reducing agent is oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

26. The composition of any one of claims 8-10 and 21-25, wherein said lysis buffer comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10- Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l ,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'- tetraacetic acid (BAPTA), tetrahydropyran (THP), or any combination thereof.

27. The composition of claim 26, wherein a final concentration of EGTA in said lysis buffer in the presence of a sample is about 0.1 millimolar (mM) to 10 mM, a final concentration of EDTA in said lysis buffer in the presence of a sample is about 0.1 mM to 5 mM, a final concentration of TCEP in said lysis buffer in the presence of a sample is about 1 mM to 20 mM, or a final concentration of Tris in said lysis buffer in the presence of a sample is about 1 mM to 60 mM.

28. The composition of any one of claims 1-27, wherein said composition further comprises an agent capable of reducing a disulfide bond.

29. The composition of claim 28, wherein said agent capable of reducing said disulfide bond comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2- mercaptoehtanol (0ME).

30. The composition of any one of claims 1-29, wherein said detergent is present in said composition mixed with a sample at a final concentration that is effective for lysing cells.

31. The composition of any one of claims 1-30, wherein said cyclodextrin is present in said composition mixed with a sample at a final concentration that is effective for isolating said detergent within said composition.

32. The composition of any one of claims 1-31, wherein said detergent is configured to form a complex with said solubilizer and / or said cyclodextrin to stabilize said enzyme.

33. The composition of claim 32, wherein said cyclodextrin is configured to increase the efficiency of forming said complex.

34. The composition of any one of claims 1-33, wherein said cyclodextrin has a higher binding affinity toward said detergent than a binding affinity of said solubilizer.

35. The composition of any one of claims 30-34, wherein said final concentration of said detergent is about 0.1% to 10% w / v (g of solute / 100 mL of solution).

36. The composition of any one of claims 31-35, wherein said final concentration of said cyclodextrin is about 0.1 mM to 70 mM.

37. The composition of any one of claims 1-36, wherein said cyclodextrin comprises hydroxypropyl p-cyclodextrin, hydroxypropyl y-cyclodextrin, (2-hydroxypropyl)-a- cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P-cyclodextrin, 2,6-Di- O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy- (2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, an anionic cyclodextrin, or any combination thereof.

38. The composition of any one of claims 1-37, wherein said solubilizer is present in said composition mixed with a sample at a final concentration of about 0.1% to 50% w / v.

39. The composition of claim 38, wherein said final concentration of said solubilizer is effective for forming micelles comprising said detergent.

40. The composition of claim 9 or 10, wherein said recovery buffer comprises a salt.

41. The composition of claim 9 or 10, wherein said recovery buffer does not comprise a salt.

42. The composition of any one of claims 9, 10, 40 and 41, wherein said recovery buffer comprises a pH buffer.

43. The composition of any one of claims 9, 10, 40 and 41, wherein said recovery buffer does not comprise a pH buffer.

44. The composition of any one of claims 8-10 and 21-39, wherein said lysis buffer is lyophilized.

45. The composition of any one of claims 9, 10 and 40-43, wherein said recovery buffer is lyophilized.

46. The composition of any one of claims 1-45, wherein said composition further comprises a sample.

47. The composition of claim 46, wherein said sample is a biological sample.

48. The composition of claim 47, wherein said biological sample comprises a target nucleic acid molecule subject to sample processing.

49. The composition of any one of claims 1-48, wherein said composition further comprises a reaction mixture for nucleic acid amplification.

50. The composition of claim 49, wherein said reaction mixture is lyophilized.

51. The composition of claim 49 or 50, wherein said reaction mixture comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, or a probe.

52. The composition of claim 51, wherein said composition is configured to stabilize enzymatic activity of said thermostable enzyme for use during a nucleic acid amplification.

53. The composition of claim 52, wherein said thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a B st 2.0 polymerase, a B st 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and any mutants thereof.

54. The composition of claim 51, wherein said dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP.

55. The composition of claim 54, wherein a concentration of said dNTPs in said reaction mixture is about 40 micromolar (pM) to 5000 pM.

56. The composition of claim 51, wherein said primer is at least 4 nucleotides in length.

57. A method of processing a sample, said method comprising mixing a sample with said lysis buffer of any one of claims 8-10 or 21-56.

58. The method of claim 57, further comprising mixing said sample with said recovery buffer of any one of claims 9, 10, or 40-56.

59. A method of processing a sample, the method comprising:(a) contacting said sample with a lysis buffer comprising a detergent; and(b) contacting said sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing said sample to generate a processed sample in a mixture comprising said detergent, said solubilizer, and said cyclodextrin.-MO-60. The method of claim 59, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

61. The method of claim 59 or 60, wherein said solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

62. The method of any one of claims 59-61, wherein said sample is a biological sample.

63. The method of any one of claims 59-62, wherein said sample is a purified sample.

64. The method of any one of claims 59-63, wherein (a) and (b) occur simultaneously.

65. The method of any one of claims 59-64, wherein contacting said sample in (a) and (b) is performed concurrently in the same mixture.

66. The method of any one of claims 59-65, further comprising incubating said sample at room temperature for a duration of time.

67. The method of any one of claims 59-66, further comprising heating said sample at a constant temperature for a period of time.

68. The method of any one of claims 59-66, further comprising heating said sample at a cyclic temperature for a period of time.

69. The method of claim any one of claims 59-68, further comprising sonicating said sample.

70. The method of any one of claims 67 or 69, wherein sonicating said sample occurs prior to, subsequent to, or concurrent to heating said sample.

71. A method of processing a sample, the method comprising:(a) contacting said sample with a lysis buffer comprising a detergent;(b) incubating said sample at a first temperature or temperature range for a first time period;(c) heating said sample at a second temperature or temperature range for a second time period; and(d) contacting said sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing said sample to generate a processed sample in a mixture comprising said detergent, said solubilizer and said cyclodextrin.

72. The method of claim 71, wherein the heating said sample in (c) further comprises heating said sample to said second temperature, cooling down said sample, and heating said sample to said second temperature after cooling down.

73. The method of claim 71 or 72, further comprising sonicating said sample.

74. The method of claim 73, wherein said sonicating said sample is performed prior to, subsequent to, or concurrent to heating said sample.

75. The method of any one of claims 71-74, further comprising bead beating said sample.

76. The method of any one of claims 71-75, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

77. The method of any one of claims 71-76, wherein said solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

78. The method of any one of claims 71-77, wherein said sample is a biological sample.

79. The method of any one of claims 71-78, wherein said sample is a purified sample.

80. The method of any one of claims 71-79, wherein said first temperature or temperature range is about 2°C to 25°C.

81. The method of any one of claims 71-80, wherein said second temperature is about 60°C to 100°C.

82. The method of any one of claims 71-81, wherein said first time period is at least about 1 minute to at least about 48 hours.

83. The method of any one of claims 71-82, wherein said second time period is at least about 1 minute to at least about 10 minutes.

84. The method of any one of claims 59-83, wherein said lysis buffer further comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2- carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10- Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l ,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'- tetraacetic acid (BAPTA), or tetrahydropyran (TEIP).

85. The method of any one of claims 59-84, further comprising, subsequent to contacting said sample with said recovery buffer, contacting said processed sample with a reaction mixture.

86. The method of claim 85, wherein said reaction mixture comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, or a probe.

87. The method of claim 86, wherein said reaction mixture stabilizes enzymatic activity of said thermostable enzyme for use during a nucleic acid amplification.

88. The method of claim 87, wherein said thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo- Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and any mutants thereof.

89. The method of claim 86, wherein said dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP.

90. The method of claim 89, wherein a concentration of said dNTPs in said reaction mixture is about 40 pM to 5000 pM.

91. The method of claim 86, wherein said primer is at least 4 nucleotides in length.

92. The method of any one of claims 85-91, further comprising subjecting said processed sample mixed with said reaction mixture to a nucleic acid amplification.

93. The method of claim 92, wherein said nucleic acid amplification comprises polymerase chain reaction (PCR) or isothermal amplification.

94. The method of claim 92 or 93, wherein said nucleic acid amplification comprises thermocycling said processed sample.

95. The method of any one of claims 92-94, wherein said nucleic acid amplification comprises keeping said processed sample at a constant temperature for amplification.

96. The method of any one of claims 59-95, wherein said method further comprising, prior to (a), obtaining said sample from a subject.

97. The method of claim 96, wherein said subject has or is suspected of having a disease, a condition, or an infection.

98. The method of any one of claims 59-97, wherein said sample comprises one or more different target nucleic acid molecules.

99. The method of any one of claims 59-98, wherein said sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, a lymph sample, raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof.

100. The method of any one of claims 96-99, wherein a time from obtaining said sample to generating said processed sample is equal to or less than about 30 min, 25 min, 20 min, 15 min, 10 min, 5 min, 4 min, 3 min, 2 min, 1 min or less.

101. The method of any one of claims 98-100, wherein a concentration of said one or more different target nucleic acid molecules is at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50% or more compared to a concentration of one or more different target nucleic acid molecules of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually.

102. A kit for sample processing, the kit comprising a lysis buffer comprising a detergent, a recovery buffer comprising a solubilizer and a cyclodextrin, and an instruction for use.

103. The kit of claim 102, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

104. The kit of claim 102 or 103, wherein said solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

105. The kit of any one of claims 102-104, further comprising a reagent for nucleic acid amplification comprising a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), or a primer.

106. The kit of claim 105, wherein said thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo- Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exo-polymerase, an OmniTaq 2 LA DNA polymerase, and any mutants thereof.

107. The kit of claim 105, wherein said dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP.

108. The kit of claim 105 or 107, wherein a concentration of said dNTPs in a reaction mixture is about 40 pM to 5000 pM.

109. The kit of claim 105, wherein said primer is at least 4 nucleotides in length.

110. The kit of any one of claims 102-109, wherein said kit further comprises a probe for detecting an amplification product generated using said kit.

111. The kit of any one of claims 102-110, wherein said lysis buffer, said recovery buffer or said reagent is lyophilized.

112. The composition of claim 9, wherein said recovery buffer further comprises a cucurbituril.

113. The composition of claim 112, wherein the cucurbituril is cucurbit[n]uril, wherein n is an integer of 5, 6, 7, 8, or 10.

114. The composition of any one of claims 49-56, wherein said reaction mixture comprises an excipient.

115. The composition of claim 114, wherein said excipient comprises one or more reagents selected from the group consisting of a Tris, potassium phosphate, sodium chloride, ethylenediaminetetraacetic acid (EDTA), potassium chloride, nonoxynol-9, trehalose, dextran, polysucrose 400, and a cyclodextrin.

116. The composition of claim 115, wherein said cyclodextrin comprises hydroxypropyl 0- cyclodextrin, hydroxypropyl y-cyclodextrin, (2-hydroxypropyl)-a-cyclodextrin, 3A- amino-3A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-0- cyclodextrin, 6-O-alpha-D-Maltosyl-p-cyclodextrin, 2,6-Di-O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, an anionic cyclodextrin, or any combination thereof.

117. The composition of claim 115 or 116, wherein: a final concentration of Tris in said excipient in the presence of said sample is about 0.001 molar (M) to 1.0 M; a final concentration of sodium chloride and / or potassium chloride in the presence of said sample is about 0.0001 M to 0.25 M; a final concentration of EDTA in said excipient in the presence of said sample is about 0.00001 M to 0.1 M; a final concentration of nonoxynol-9 in said excipient in the presence of said sample is about 0.01% v / v to 2.0% v / v; a final concentration of trehalose in said excipient in the presence of said sample is about .001 M to 2.0 M; a final concentration of dextran in said excipient in the presence of said sample is about 0.1% w / v to 10% w / v; a final concentration of poly sucrose 400 in said excipient in the presence of said sample is about 0.01% w / v to 5.0% w / v; and / ora final concentration of said cyclodextrin in said excipient in the presence of said sample is about 0.001 M to 5.0 M.

118. The composition of any one of claims 114-117, wherein said excipient further comprises an additional reagent.

119. The composition of claim 118, wherein said additional reagent comprises a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof.

120. The composition of claim 118 or 119, wherein said additional reagent is configured to stabilize an enzyme.

121. The composition of any one of claims 118-120, wherein said additional reagent is configured to lower a Cq value of a nucleic acid amplification.

122. The composition of any one of claims 49-56 and 114-121, wherein said composition further comprises a sample stabilization buffer.

123. The composition of claim 122, wherein said sample stabilization buffer comprises one or more reagents selected from the group consisting of a collapse modifier, a protein stabilizer, and a glass transition modifier.

124. The composition of claim 122 or 123, wherein said sample stabilization buffer comprises at least one salt.

125. The composition of any one of claims 122-124, wherein said sample stabilization buffer comprises a cyclodextrin.

126. The composition of any one of claims 122-125, wherein said sample stabilization buffer is configured to reconstitute a lyophilized sample.

127. The composition of any one of claims 122-126, wherein said stabilization buffer comprises at least one reducing agent.

128. The composition of claim 127, wherein said at least one reducing agent is oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, 1 ,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP), or any combination thereof.

129. The method of any one of claims 59-70, wherein a total time to perform (a) and (b) is at most 1 min, at most 50 seconds, at most 40 seconds, or at most 20 seconds.

130. The method of any one of claims 85-95, wherein a time for processing said sample is a time period from said contacting of (a) to contacting said processed sample with a reaction mixture, wherein said time period is at most 20 seconds.

131. The method of claim 92, wherein said nucleic acid amplification generates an amplified processed sample.

132. The method of claim 131, wherein a time period of said nucleic acid amplification to generate said amplified processed sample is at most 5 minutes.

133. The method of any one of claims 59-101 and 129-132, wherein said method of processing said sample does not comprise heating said sample.

134. A composition for sample amplification comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer, wherein said composition is configured to increase a rate of amplification during a nucleic acid amplification.

135. The composition of claim 134, wherein said composition is configured to stabilize an enzyme during a nucleic acid amplification.

136. The composition of claim 135, wherein said enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof.

137. The composition of claim 136, wherein said reverse transcriptase is an avian myeloblastosis virus (AMV) reverse transcriptase or a murine leukemia virus (MMLV) reverse transcriptase.

138. The composition of any one of claims 134-136, wherein said nonionic surfactant is nonoxynol-9.

139. The composition of any one of claims 134-138, wherein a final concentration of said cyclodextrin in said composition in the presence of a sample is 0.01% v / v to 2.0% v / v.

140. The composition of any one of claims 134-139, wherein said cyclodextrin comprises hydroxypropyl P-cyclodextrin, hydroxypropyl y-cyclodextrin, (2-hydroxypropyl)-a- cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P-cyclodextrin, 2,6-Di- O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy- (2AS,3AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, or any combination thereof.

141. The composition of any one of claims 134-140, wherein a final concentration of said cyclodextrin in said composition in the presence of a sample is about 0.001 M to 10 M.

142. The composition of any one of claims 134-141, wherein said sucrose / epichlorohydrin polymer is polysucrose 400.

143. The composition of any one of claims 134-142, wherein a final concentration of said sucrose / epichlorohydrin polymer in said composition in the presence of a sample is about 0.001% to 5% w / v (g of solute / 100 mL of solution).

144. The composition of any one of claims 134-143, wherein said composition further comprises at least one salt.

145. The composition of claim 144, wherein a final concentration of said at least one salt in said composition in the presence of a sample is about 0.001 molar (M) to 10 M.

146. The composition of claim 144 or 145, wherein said at least one salt is sodium chloride, potassium phosphate, potassium chloride, or any combination thereof.

147. The composition of any one of claims 134-146, wherein said composition comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2- carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10- Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l ,2-ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'- tetraacetic acid (BAPTA), or tetrahydropyran (TEIP).

148. The composition of claim 147, wherein:(a) a final concentration of EDTA in said composition in the presence of a sample is about 0.01 millimolar (mM) to 10 mM, and / or(b) a final concentration of Tris in said composition in the presence of a sample is about 0.1 mM to 25 mM.

149. The composition of any one of claims 134-148, wherein said composition further comprises an agent capable of reducing a disulfide bond.

150. The composition of claim 149, wherein said agent capable of reducing said disulfide bond comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2- mercaptoehtanol (0ME).

151. The composition of any one of claims 134-150, wherein said composition further comprises at least one sugar and / or sugar alcohol.

152. The composition of claim 151, wherein said at least one sugar and / or sugar alcohol comprises sucrose, lactose, trehalose, dextran, erythritol, arabitol, xylitol, sorbitol, mannitol, or any combination thereof.

153. The composition of claim 151 or 152, wherein a final concentration of said at least one sugar and / or sugar alcohol is about 0.001 M to 10 M or about 0.1% to 10% w / v (g of solute / 100 mL of solution).

154. The composition of any one of claims 134-153, wherein said composition further comprises an additional reagent.

155. The composition of claim 154, wherein said additional reagent comprises a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof.

156. The composition of any one of claims 134-155, wherein said composition is lyophilized.

157. The composition of any one of claims 134-156, wherein said composition further comprises a sample.

158. The composition of claim 157, wherein said sample is a biological sample.

159. The composition of claim 158, wherein said biological sample comprises a target nucleic acid molecule subject to sample processing.

160. The composition of any one of claims 134-159, wherein said composition further comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, a probe, or any combination thereof.

161. The composition of claim 160, wherein said composition is configured to stabilize enzymatic activity of said thermostable enzyme for use during said nucleic acid amplification.

162. The composition of claim 160 or 161, wherein said thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exopolymerase, an OmniTaq 2 LA DNA polymerase, a IsoFast™ Bst, and any mutants thereof.

163. The composition of any one of claims 160-162, wherein said dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP.

164. The composition of claim 163, wherein a concentration of said dNTPs in said composition is about 40 micromolar (pM) to 5000 pM.

165. The composition of any one of claims 160-164, wherein said primer is at least 4 nucleotides in length.

166. The composition of any one of claims 160-165, wherein said probe is at least 15 nucleotides in length.

167. A composition comprising: a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer; and a sample stabilization buffer configured to stabilize an enzyme in a nucleic acid amplification.

168. The composition of claim 167, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

169. The composition of claim 167 or 168, wherein said solubilizer is a non-ionic surfactant.

170. The composition of any one of claims 167-169, wherein said solubilizer is a polysorbate, octylphenoxypolyethoxyethanol, 2-[4-(2,4,4-trimethylpentan-2- yl)phenoxy]ethanol, or a secondary alcohol ethoxylate.

171. The composition of claim 170, wherein said solubilizer is polysorbate 80, polysorbate 20, polysorbate 40, polysorbate 60, or a functional variant thereof.

172. The composition of any one of claims 167-171, wherein said detergent is part of a lysis buffer.

173. The composition of any one of claims 167-172, wherein said solubilizer and said cyclodextrin are part of a recovery buffer.

174. The composition of claim 173, wherein said lysis buffer and said recovery buffer are in said sample processing buffer as the same mixture.

175. The composition of any one of claims 167-174, wherein said solubilizer and said cyclodextrin are configured to shorten a cycle threshold value or a time to result value in said nucleic acid amplification compared to a cycle threshold value or a time to result value in a nucleic acid amplification of an otherwise identical sample processed by SDS, polysorbate 80, or a cyclodextrin individually.

176. The composition of claim 175, wherein said cycle threshold value is at most 40 or said time to result value is at most 15 minutes.

177. The composition of any one of claims 167-176, wherein said solubilizer and said cyclodextrin are configured to decrease a coefficient of variation.-ISO-178. The composition of any one of claims 167-177, wherein said solubilizer and said cyclodextrin are configured to lower a limit of detection.

179. The composition of claim 172 or 174, wherein said lysis buffer further comprises a chelating agent.

180. The composition of claim 179, wherein said chelating agent is deferiprone, ethylenediamine, 1,10-Phenanthroline, oxalic acid, pentetic acid, deferasirox, deferoxamine, deferoxamine mesylate, or N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2- ethanediamine (TPEN).

181. The composition of any one of claims 172, 174, and 179-180, wherein said lysis buffer further comprises a reducing agent.

182. The composition of claim 181, wherein said reducing agent is oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, 1 ,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

183. The composition of any one of claims 172, 174, and 179-182, wherein said lysis buffer comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, a ethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2- ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

184. The composition of claim 183, wherein a final concentration of EGTA in said lysis buffer in the presence of a sample is aboutO.1 millimolar (mM) to 10 mM, a final concentration of EDTA in said lysis buffer in the presence of a sample is about 0.1 mM to 5 mM, a final concentration of TCEP in said lysis buffer in the presence of a sample is about 1 mM to 20 mM, or a final concentration of Tris in said lysis buffer in the presence of a sample is about 1 mM to 60 mM.

185. The composition of any one of claims 167-184, wherein said sample processing buffer further comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2-mercaptoehtanol ( ME).

186. The composition of any one of claims 167-185, wherein said detergent is present in said sample processing buffer mixed with a sample at a final concentration that is effective for lysing cells.

187. The composition of any one of claims 167-186, wherein said cyclodextrin is present in said sample processing buffer mixed with a sample at a final concentration that is effective for isolating said detergent within said composition.

188. The composition of any one of claims 167-187, wherein said detergent is configured to form a complex with said solubilizer and / or said cyclodextrin to stabilize said enzyme.

189. The composition of claim 188, wherein said cyclodextrin is configured to increase the efficiency of forming said complex.

190. The composition of claim 186, wherein said final concentration of said detergent is about 0.1% to 10% w / v (g of solute / 100 mL of solution).

191. The composition of claim 187, wherein said final concentration of said cyclodextrin is about 0.1 mM to 70 mM.

192. The composition of any one of claims 167-191, wherein said cyclodextrin comprises hydroxypropyl P-cyclodextrin, hydroxypropyl y-cyclodextrin, (2-hydroxypropyl)-a- cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-P-cyclodextrin, 6-O-alpha-D-Maltosyl-P-cyclodextrin, 2,6-Di- O-methyl-P-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy- (2AS, 3 AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, or any combination thereof.

193. The composition of any one of claims 167-192, wherein said solubilizer is present in said composition mixed with a sample at a final concentration of about 0.1% to 50% w / v.

194. The composition of claim 193, wherein said final concentration of said solubilizer is effective for forming micelles comprising said detergent.

195. The composition of claim 173 or 174, wherein said recovery buffer comprises a salt.

196. The composition of claim 195, wherein said recovery buffer comprises a pH buffer.

197. The composition of claim 195, wherein said recovery buffer does not comprise a pH buffer.

198. The composition of any one of claims 167-197, wherein said sample processing buffer is lyophilized.

199. The composition of any one of claims 167-198, wherein said nonionic surfactant of said sample amplification buffer is nonoxynol-9.

200. The composition of any one of claims 167-199, wherein a final concentration of said cyclodextrin in said sample amplification buffer in the presence of a sample is 0.01% v / v to 2.0% v / v.

201. The composition of claim 200, wherein said cyclodextrin comprises said cyclodextrin comprises hydroxypropyl p-cyclodextrin, hydroxypropyl y-cyclodextrin, (2- hydroxypropyl)-a-cyclodextrin, 3A-amino-3A-deoxy-(2AS,3AS)-a-cyclodextrin hydrate, monopropanediamino-p-cyclodextrin, 6-O-alpha-D-Maltosyl-P-cyclodextrin, 2,6-Di-O-methyl-p-cyclodextrin, hydroxyethyl-P-cyclodextrin, 3A-amino-3A-deoxy- (2AS, 3 AS)-P-cyclodextrin hydrate, 3A-amino-3A-deoxy-(2AS,3AS)-y-cyclodextrin hydrate, or any combination thereof.

202. The composition of claim 200 or 201, wherein a final concentration of said cyclodextrin in said sample amplification buffer in the presence of a sample is about 0.001 M to 10 M.

203. The composition of any one of claims 167-202, wherein said sucrose / epichlorohydrin polymer is polysucrose 400.

204. The composition of claim 203, wherein a final concentration of said sucrose / epichlorohydrin polymer in said composition in the presence of a sample is about 0.001% to 5% w / v (g of solute / 100 mL of solution).

205. The composition of any one of claims 167-204, wherein said sample amplification buffer further comprises at least one salt.

206. The composition of claim 205, wherein a final concentration of said at least one salt in said composition in the presence of a sample is about 0.001 molar (M) to 10 M.

207. The composition of claim 205 or 206, wherein said at least one salt is sodium chloride, potassium phosphate, potassium chloride, or any combination thereof.

208. The composition of any one of claims 167-207, wherein said sample amplification buffer further comprises an egtazic acid (EGTA), an ethylenediaminetetraacetic acid (EDTA), a tris(2-carboxyethyl)phosphine (TCEP), a Tris, a deferiprone, aethylenediamine, 1,10-Phenanthroline, an oxalic acid, a pentetic acid, a deferasirox, a deferoxamine, a deferoxamine mesylate, N,N,N',N'-tetrakis(2-pyridinylmethyl)-l,2- ethanediamine (TPEN), a formic acid, a lithium aluminum hydride, a sodium borohydride, a thiosulfate, a sodium hydrosulfite, l,2-bis(o-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP).

209. The composition of claim 208, wherein:(a) a final concentration of EDTA in said composition in the presence of a sample is about 0.01 millimolar (mM) to 10 mM, and / or(b) a final concentration of Tris in said composition in the presence of a sample is about 0.1 mM to 60 mM.

210. The composition of any one of claims 167-209, wherein said sample amplification buffer further comprises an agent capable of reducing a disulfide bond.

211. The composition of claim 210, wherein said agent capable of reducing said disulfide bond comprises dithiothreitol (DTT), tris(2-carboxyethyl)phosphine (TCEP), or 2- mercaptoehtanol (0ME).

212. The composition of any one of claims 167-211, wherein said sample amplification buffer further comprises at least one sugar and / or sugar alcohol.

213. The composition of claim 212, wherein said at least one sugar and / or sugar alcohol comprises sucrose, lactose, trehalose, dextran, erythritol, arabitol, xylitol, sorbitol, mannitol, or any combination thereof.

214. The composition of claim 212 or 213, wherein a final concentration of said at least one sugar and / or sugar alcohol is about 0.001 M to 10 M or about 0.1% to 10% w / v (g of solute / 100 mL of solution).

215. The composition of any one of claims 167-214, wherein said sample amplification buffer further comprises an additional reagent.

216. The composition of claim 215, wherein said additional reagent comprises a base, Brij 98, guanidinium thiocyanate (GITC), methionine, non-detergent sulfobetaine (NDSB), tRNA, recombinant Albumin (rAlbumin), or any combination thereof.

217. The composition of any one of claims 167-216, wherein said sample amplification buffer is lyophilized.

218. The composition of any one of claims 167-217, wherein said sample amplification buffer further comprises a thermostable enzyme, deoxynucleoside triphosphates (dNTPs), a primer, a probe, or any combination thereof.

219. The composition of claim 218, wherein said sample amplification buffer is configured to stabilize enzymatic activity of said thermostable enzyme for use during said nucleic acid amplification.

220. The composition of claim 218 or 219, wherein said thermostable enzyme is selected from the group consisting of a large fragment of a Bacillus stearothermophilus polymerase, a exo-Klenow polymerase, a Bst 2.0 polymerase, a Bst 3.0 polymerase, a SD DNA polymerase, a phi29 DNA polymerase, a sequencing-grade T7 exopolymerase, an OmniTaq 2 LA DNA polymerase, a IsoFast™ Bst, and any mutants thereof.

221. The composition of any one of claims 218-220, wherein said dNTPs comprise dATP, dCTP, dGTP, dTTP, or dUTP.

222. The composition of claim 221, wherein a concentration of said dNTPs in said composition is about 40 micromolar (pM) to 5000 pM.

223. The composition of any one of claims 218-222, wherein said primer is at least 4 nucleotides in length.

224. The composition of any one of claims 218-223, wherein said probe is at least 15 nucleotides in length.

225. The composition of any one of claims 167-224, wherein said sample stabilization buffer comprises one or more reagents selected from the group consisting of a collapse modifier, a protein stabilizer, and a glass transition modifier.

226. The composition of any one of claims 167-225, wherein said sample stabilization buffer comprises at least one salt.

227. The composition of any one of claims 167-226, wherein said sample stabilization buffer comprises a cyclodextrin.

228. The composition of any one of claims 167-227, wherein said sample stabilization buffer is configured to reconstitute a lyophilized sample.

229. The composition of any one of claims 167-228, wherein said stabilization buffer comprises at least one reducing agent.

230. The composition of claim 229, wherein said at least one reducing agent is oxalic acid, formic acid, lithium aluminum hydride, sodium borohydride, a thiosulfate, sodium hydrosulfite, 1 ,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), or tetrahydropyran (THP), or any combination thereof.

231. The composition of any one of claims 167-230, wherein said composition further comprises a sample.

232. The composition of claim 231, wherein said sample is a biological sample.

233. The composition of claim 232, wherein said biological sample comprises a target nucleic acid molecule subject to sample processing.

234. The composition of any one of claims 167-233, wherein said enzyme is a polymerase, an endonuclease, a reverse transcriptase, or any combination thereof.

235. A method of amplifying a sample, the method comprising:(a) contacting said sample with a sample processing buffer to generate a processed sample;(b) contacting said processed sample with a sample amplification buffer to provide a condition for a nucleic acid amplification; and(c) subjecting said processed sample to said nucleic acid amplification, and wherein prior to said contacting of (b), said sample processing buffer is not removed.

236. The method of 235, further comprising contacting said sample with a sample stabilization buffer for stabilizing an enzyme in said nucleic acid amplification.

237. The method of claim 236, further comprising, prior to (c), contacting said sample with said sample stabilization buffer.

238. The method of claim 236 or 237, wherein said sample stabilization buffer is in a same mixture as said sample amplification buffer.

239. The method of any one of claims 236-238, wherein said sample stabilization buffer is contacted with said sample after contacting said sample with said sample amplification buffer.

240. The method of any one of claims 235-239, wherein said method does not comprise heating said sample.

241. The method of any one of claims 235-240, wherein said sample processing buffer comprises a lysis buffer and / or a recovery buffer.

242. The method of claim 241, wherein said lysis buffer comprises a detergent.

243. The method of claim 242, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

244. The method of any one of claims 241-243, wherein said recovery buffer comprises a solubilizer and a cyclodextrin.

245. The method of any one of claims 235-244, wherein a processing time for said sample is a time period from said contacting of (a) and said contacting of (b), wherein said time period is at most 20 seconds.

246. The method of any one of claims 235-245, wherein said nucleic acid amplification comprises polymerase chain reaction (PCR) or isothermal amplification.

247. The method of any one of claims 235-246, wherein said nucleic acid amplification comprises thermocycling said sample.

248. The method of any one of claims 235-247, wherein said nucleic acid amplification generates an amplified sample.

249. The method of claim 248, wherein a time period from said contacting of (a) to said amplified sample is at most 5 minutes.

250. The method of any one of claims 235-249, wherein said sample is a biological sample.

251. The method of claim 250, wherein said biological sample comprises one or more different target nucleic acid molecules.

252. The method of claim 251, wherein a concentration of said one or more different target nucleic acid molecules is at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% greater than a concentration of one or more different target nucleic acid molecules of an otherwise identical sample processed by a sample processing buffer alone.

253. The method of any one of claims 235-252, wherein the sample processing buffer further comprises a cucurbituril.

254. A method of processing a sample, the method comprising:(a) contacting said sample with a sample processing buffer to generate a processed sample;(b) contacting said processed sample with a sample amplification buffer to provide a condition for a nucleic acid amplification; and(c) subjecting said processed sample to said nucleic acid amplification in said sample amplification buffer, wherein a time period from contacting in (a) to generating said processed sample prior to contacting with said sample amplification buffer is (i) no more than a time for pipetting said sample processing buffer into said sample to mix said sampleprocessing buffer and said sample or is (ii) at most 1 min, at most 50 seconds, at most 40 seconds, or at most 20 seconds.

255. The method of claim 254, wherein said sample processing buffer comprises a lysis buffer and / or a recovery buffer.

256. The method of claim 255, wherein said lysis buffer comprises a detergent.

257. The method of claim 256, wherein said detergent is sodium dodecyl sulfate (SDS), sodium lauryl sulfate, lithium dodecyl sulfate, or a functional variant thereof.

258. The method of any one of claims 255-257, wherein said recovery buffer comprises a solubilizer and a cyclodextrin.

259. The method of any one of claims 254-258, wherein said sample processing buffer comprises a detergent, a solubilizer, and a cyclodextrin, wherein said sample processing buffer is configured to stabilize an enzyme during a nucleic acid amplification, and wherein said sample processing buffer is configured to reduce and / or eliminate activity of a degrading nuclease.

260. The method of any one of claims 254-259, wherein said sample processing buffer is the composition of any one of claims 1-48.

261. The method of any one of claims 254-260, wherein said sample amplification buffer comprises an excipient.

262. The method of any one of claims 254-261, wherein, prior to (b), said sample processing buffer is not removed.

263. The method of any one of claims 254-262, wherein said sample amplification buffer comprises said reaction mixture of any one of claims 114-128, or said sample amplification buffer is the composition of any one of claims 134-164.

264. The method of any one of claims 254-263, wherein said method does not comprise heating said sample.

265. The method of any one of claims 254-264, wherein said sample is a biological sample.

266. The method of claim 265, wherein said biological sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, a lymph sample, raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, anenvironmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof.

267. The method of claim 266, wherein said blood sample is obtained from a subject.

268. The method of claim 266 or 267, wherein said blood sample is collected in a blood collection tube.

269. The method of claim 268, wherein said blood collection tube comprises a stabilizing agent for stabilizing RNAs.

270. The method of claim 269, wherein said stabilizing agent comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid.

271. The method of claim 269 or 270, wherein said blood sample is contacted with said sample processing buffer without removing said stabilizing agent.

272. The method of any one of claims 266-271, wherein said blood sample is contacted with said sample processing buffer directly without being subject to other processing prior to contacting said sample processing buffer.

273. The method of claim 271 or 272, wherein said blood sample is not processed by centrifugation or a spin column prior to contacting said sample processing buffer.

274. The method of any one of claims 254-273, wherein said sample is lyophilized.

275. The method of any one of claims 254-274, wherein said sample amplification buffer is lyophilized.

276. The method of any one of claims 254-275, wherein the sample processing buffer further comprises a cucurbituril.

277. A composition for sample processing comprising: a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin; a stabilizing agent comprising tetradecyl trimethyl-ammonium oxalate and / or tartaric acid, and wherein said composition is configured to stabilize an enzyme during a nucleic acid amplification, and wherein said composition is configured to reduce and / or eliminate activity of a degrading nuclease.

278. A composition for sample processing comprising: a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer;a stabilizing agent comprising tetradecyl trimethyl-ammonium oxalate and / or tartaric acid, and wherein said composition is configured to increase a rate of amplification during a nucleic acid amplification.

279. The composition of claim 277 or 278, wherein said composition does not comprise ethanol.

280. The composition of any one of claims 277-279, wherein the composition further comprises cucurbituril.

281. The composition of claim 280, wherein the cucurbituril comprises cucurbit[n]uril, wherein n is an integer of 5, 6, 7, 8, or 10.

282. The composition of claim 280, wherein the cucurbituril is cucurbit[7]uril.

283. The composition of any one of claims 277-282, wherein the composition further comprises a sample.

284. The composition of claim 283, wherein said sample is a biological sample.

285. The composition of claim 284, wherein said biological sample comprises a blood sample, a swab sample, a saliva sample, a urine sample, a cerebrospinal fluid sample, a pleural fluid sample, a rectal sample, a vaginal sample, a stool sample, a sputum sample, a lymph sample, raw milk, pasteurized and / or homogenized milk, pasteurized and / or processed milk, one or more Bacillus anthracis spores, one or more Bacillus anthracis vegetative cells, a tissue sample, a cell culture, a purified nucleic acid sample, an environmental sample, one or more whole organisms, one or more homogenized organisms, wastewater, or any combination thereof.

286. The composition of claim 285, wherein said blood sample is obtained from a subject.

287. The composition of claim 285 or 286, wherein said blood sample is collected in a blood collection tube.

288. The composition of claim 287, wherein said blood collection tube comprises a stabilizing agent for stabilizing RNAs.

289. The composition of claim 288, wherein said stabilizing agent comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid.

290. The composition of claim 288 or 289, wherein said blood sample is contacted with said sample processing buffer without removing said stabilizing agent.

291. The composition of any one of claims 285-290, wherein said blood sample is contacted with said sample processing buffer directly without being subject to other processing prior to contacting said sample processing buffer.

292. The composition of claim 290 or 291, wherein said blood sample is not processed by centrifugation or a spin column prior to contacting said processing buffer.

293. A method of processing a sample, the method comprising:(a) contacting said sample with a lysis buffer comprising a detergent, wherein said sample comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid or wherein said sample is directly from a sample collection tube; and / or(b) contacting said sample with a recovery buffer comprising a solubilizer and a cyclodextrin, thereby processing said sample to generate a processed sample in a mixture comprising said detergent, said solubilizer, and said cyclodextrin.

294. The method of claim 293, further comprising contacting said sample with a sample amplification buffer.

295. A method of processing a sample, the method comprising: contacting said sample with a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin, and a sucrose / epichlorohydrin polymer, wherein said composition is configured to increase a rate of amplification during a nucleic acid amplification, and wherein said sample comprises tetradecyl trimethyl- ammonium oxalate and / or tartaric acid or wherein said sample is directly from a sample collection tube.

296. The method of claim 295, further comprising, prior to contacting said sample with said sample amplification buffer, contacting sample with a sample processing buffer.

297. A method of processing a sample, the method comprising:(a) contacting said sample with a sample processing buffer comprising: a detergent, a solubilizer, and a cyclodextrin;(b) contacting said sample with a sample amplification buffer comprising: a nonionic surfactant, a cyclodextrin; and(c) contacting said sample with a sample stabilization buffer configured to stabilize an enzyme in a nucleic acid amplification, wherein said sample comprises tetradecyl trimethyl-ammonium oxalate and / or tartaric acid or wherein said sample is directly from a sample collection tube.

298. The method of any one of claims 293-297, wherein said sample is not processed by an RNA extraction kit.

299. The method of claim 298, wherein said kit comprises a spin-column.

300. The method of claim 298 or 299, wherein said kit comprises a wash pellet.

301. The method of any one of claims 293-300, wherein said method does not comprise contacting with a wash buffer.

302. The method of any one of claims 293-301, wherein said method does not comprise membrane-based extraction.

303. The method of any one of claims 293-302, further comprising subjecting said sample to a nucleic acid amplification.

304. The method of claim 303, wherein said nucleic acid amplification comprises polymerase chain reaction (PCR) or isothermal amplification.

305. The method of claim 303 or 304, wherein said nucleic acid amplification comprises thermocycling said sample.

306. The method of any one of claims 303-305, wherein said nucleic acid amplification generates an amplified sample.

307. The method of claim 306, wherein a time period from said contacting in (a) to generating said amplified sample is at most 5 minutes.

308. The method of any one of claims 293-307, wherein a processing time for said sample is a time period from said contacting in (a) to generating a processed sample prior to contacting with said amplification buffer, wherein said processing time is at most 1 min, at most 50 seconds, at most 30 seconds, or at most 20 seconds.

309. The method of any one of claims 293-308, wherein said method does not comprise heating said sample.

310. The method of any one of claims 293-309, wherein said sample is a blood sample.

311. The method of any one of claims 293-310, further comprising obtaining said sample from a subject and collecting said sample in said sample collection tube.

312. The method of any one of claims 296-311, wherein the sample processing buffer further comprises a cucurbituril.

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