Compositions and methods for performing assay measurements

Abstract

To provide a composition and method for executing assay measurement.SOLUTION: The present disclosure provides a composition, a reagent, a kit, a system, a system component, and a method for performing assays. More specifically, the present disclosure relates to an assay composition for detecting luminescence, including an alkyldiethanolamine, e.g., N-butyldiethanolamine (BDEA). In some embodiments, the assay composition includes 2-dibutylaminoethanol (BDAE).SELECTED DRAWING: None

Description

[Technical Field]

[0001] This specification relates to compositions for use in assays, particularly electrochemiluminescent assays, and methods of using the same. See U.S. Patent Nos. 6,919,173, 7,288,410, 7,491,540, and 8,785,201, each of which is incorporated herein by reference. [Background technology]

[0002] There are many commercially available instruments that utilize electrochemiluminescence (ECL) for analytical measurements. Examples of ECL labels include: i) luminescent organometallic compounds, e.g., compounds containing lanthanide metals such as Ru, Os, Ir, Re, or tris-bipyridyl-ruthenium (RuBpy) moieties; ii) luminol and related compounds; and iii) noble metal nanoclusters, e.g., compounds of the general formula Au n (SR) mwhere n and m are integers (e.g., n and m are 18-144). The species involved in ECL labeling in the ECL process are referred to herein as ECL coreactants. Commonly used coreactants include tertiary amines (see, e.g., U.S. Pat. No. 5,846,485, incorporated herein by reference), oxalates, and persulfates for ECL from RuBpy, and hydrogen peroxide for ECL from luminol (see, e.g., U.S. Pat. No. 5,240,863). Light generated by ECL labels can be used as a reporter signal in diagnostic procedures (Bard et al., U.S. Pat. No. 5,238,808). For example, ECL labels can be covalently attached to binding agents such as antibodies, antigens, nucleic acids, receptors, ligands, or small molecules, and the participation of the binding reagent in the binding interaction can be monitored by measuring the ECL released from the ECL label. Alternatively, the ECL signal from an ECL-active compound may be an indicator of the chemical environment (see, e.g., U.S. Patent No. 5,641,623, which describes an ECL assay that monitors the formation or destruction of an ECL coreactant). For further background on ECL, ECL labels, ECL assays, and instrumentation for performing ECL assays, see U.S. Patent Nos. 5,093,268, 5,147,806, 5,324,457, 5,591,581, 5,597,910, 5,641,623, 5,643,713, 5,679,519, 5,705,402, 5,846,485, 5,866,434, 5,786,141, 5,731,147, See PCT Nos. 6,066,448, 6,136,268, 5,776,672, 5,308,754, 5,240,863, 6,207,369, 5,589,136, and published PCT Nos. WO99 / 63347, WO00 / 03233, WO99 / 58962, WO99 / 32662, WO99 / 14599, WO98 / 12539, WO97 / 36931, and WO98 / 57154.

[0003] Commercially available ECL instruments have demonstrated good performance. They have become widely used for reasons including sensitivity, dynamic range, precision, and tolerance for complex sample matrices. Several types of commercial instruments are available for performing ECL-based measurements (Debad, JD, et al., 2004. Clinical and Biological Applications of ECL, in: Electrogenerated Chemiluminescence. Marcel Dekker, pp. 43-78.). Instruments configured for performing binding assays on beads and including a flow cell for collecting beads onto an electrode for inducing ECL have been disclosed (U.S. Patent Nos. 5,935,779 and 5,993,740). ECL devices have also been disclosed that use immobilized reagents on electrodes used to induce ECL (see, e.g., U.S. Patent Nos. 6,140,045, 6,066,448, 6,090,545, 6,207,369, and published PCT application WO98 / 12539). Multiwell plates with integrated electrodes suitable for such ECL measurements have also been disclosed (see, e.g., U.S. Patent Nos. 6,977,722 and 7,842,246, which are incorporated herein by reference). Cartridges with electrodes for ECL measurements have also been disclosed (e.g., U.S. Patent Nos. 2012 / 0190589 and 2012 / 0178091).

[0004] Currently, reagents containing the ECL coreactant tripropylamine (TPA) are available to enhance light production from ECL labels. Applicants have discovered that such components have significant drawbacks. Summary of the Invention

[0005] The present disclosure relates to compositions, reagents, kits, systems, system components, and methods for performing assays. In embodiments, the disclosure relates to an assay composition for inducing luminescence, the composition comprising an alkyldiethanolamine. In embodiments, the alkyldiethanolamine has the formula (HOCH2CH2)2N-CHR 1 -CHR 2 -CHR 3 -R 4 wherein R 1 , R 2 , R 3 , and R 4 are each independently -H, -CH, -CHCH, or -CH(CH). 1 , R 2 , and R 3 is -H, and R 4 is -H or -CH3. In embodiments, the present disclosure relates to an assay composition for inducing luminescence, wherein the composition comprises N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE). In embodiments, the composition comprises BDEA.

[0006] In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a surfactant. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a liquid diluent. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, a surfactant, and a liquid diluent.

[0007] In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a surfactant. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a liquid diluent. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, a surfactant, and a liquid diluent.

[0008] In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a surfactant. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a liquid diluent. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, a surfactant, and a liquid diluent.

[0009] In some embodiments, the compositions of the present disclosure further comprise an ECL label, a binding reagent for a binding assay, a preservative, a biocide, an antifoaming agent, a perchlorate compound, a colorant, a tracer chemical, a solid support, or a combination thereof.

[0010] In some embodiments, the ECL generated by the ECL label in the presence of a composition described herein changes, on average, less than 1% per °C over a temperature range of 18 °C to 30 °C. In some embodiments, the slope of change in ECL with pH of the ECL generated by the ECL label in the presence of the composition is less than 10% per pH unit. In some embodiments, a change in the concentration of the ECL coreactant from 0.8 to 1.2 times the nominal value provides less than a 10% change in the ECL generated by the ECL label in the presence of a composition described herein. In some embodiments, the ECL is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode.

[0011] In additional embodiments, the present disclosure provides a kit comprising the following materials in one or more containers: N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both; a pH buffering component; and an ionic component.

[0012] In some embodiments, the kits of the present disclosure comprise a composition described herein and an assay device, assay consumables, additional assay reagents, an assay sample, or a combination thereof. In some embodiments, the assay device is configured to perform an ECL assay.

[0013] In a further embodiment, the present disclosure provides a method for producing a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component.

[0014] In embodiments, the present disclosure provides methods for generating ECL comprising contacting an electrode with a composition described herein and an ECL label, applying a voltage to the electrode, and generating ECL.

[0015] In embodiments, the present disclosure provides a method for measuring the amount of an ECL label, comprising contacting an electrode with a composition described herein and an ECL label, applying a voltage to the electrode, generating ECL, measuring the ECL, and determining the amount of label from the measured ECL.

[0016] In embodiments, the present disclosure provides a method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, the method comprising contacting a binding reagent immobilized on an electrode with a labeled binding reagent comprising an ECL label, forming a binding complex on the electrode comprising the immobilized binding reagent and the labeled binding reagent, contacting the binding complex on the electrode with a composition described herein, applying a voltage to the electrode in the presence of the composition, generating ECL, and measuring the ECL to determine the amount of binding complex on the electrode.

[0017] In embodiments, the present disclosure provides a method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, the method comprising contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label, forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent, contacting the binding complex on the particle with a composition described herein, collecting the particles on an electrode, applying a voltage to the electrode in the presence of the composition, generating ECL, and measuring the ECL to determine the amount of binding complex on the electrode.

[0018] In embodiments, the present disclosure provides a method for measuring the amount of an analyte, the method comprising contacting a binding reagent immobilized on an electrode with a labeled binding reagent comprising an ECL label and the analyte (or a sample comprising the analyte), forming a binding complex on the electrode comprising the immobilized binding reagent and the labeled binding reagent, contacting the binding complex on the electrode with a composition described herein, applying a voltage to the electrode in the presence of the composition, generating ECL, and measuring the ECL to determine the amount of the analyte.

[0019] In embodiments, the present disclosure provides a method for measuring the amount of an analyte, the method comprising contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label and the analyte (or a sample comprising the analyte), forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent, contacting the binding complex on the particle with a composition described herein, collecting the particles on an electrode, applying a voltage to the electrode in the presence of the composition, generating ECL, and measuring the ECL to determine the amount of the analyte. [Brief explanation of the drawings]

[0020] [Figure 1] Figure 1A shows the specific ECL signal of an ECL-labeled reagent on the electrode surface, and Figure 1B shows the background ECL signal in the absence of the labeled reagent when ECL is generated in the presence of an ECL read buffer containing BDEA, DBEA, or TPA coreactants, with or without TRITON X-100. Figure 1C also shows the effect of varying the pH buffering components in the read buffer. [Figure 2]Figures 2A and 2B show the specific and background signals generated by a multiplex panel of ECL sandwich immunoassays. Figure 2A compares the signal generated in the presence of a BDEA-containing coreactant composition with the signal generated by a conventional TPA-containing coreactant composition, and Figure 2B compares the signal generated in the presence of a BDEA-containing coreactant composition containing high levels of salt with the signal generated by a conventional TPA-containing coreactant composition. [Figure 3] Figures 3A-3F show the effects of composition and sources of environmental variation on the specific ECL signal and ECL background signal generated in the presence of an ECL read buffer containing BDEA and TPA, including the effect of read buffer pH (Figure 3A), the effect of read buffer temperature during ECL generation (Figure 3B), the effect of co-reactant concentration and the presence or absence of surfactant (Figure 3C), the effect of diluting or concentrating all components in the read buffer (Figure 3D), the effect of read buffer temperature and pH (Figure 3E), and the % background change at different read buffer temperatures and pH (Figure 3F). [Figure 4] 1 shows the effect of voltage waveform and duration on the generation of specific and background ECL signals generated in the presence of a read buffer containing BDEA and TPA. [Figure 5] FIG. 5A shows the chemical structures of a set of nonionic surfactants, and FIG. 5B shows the liquid / air interfacial tension (dynes / cm) of reading buffers prepared with these surfactants. [Figure 6] FIG. 6A shows a comparison of the specific ECL signals, and FIG. 6B shows a comparison of the ECL background signals generated using read buffers containing BDEA but with different detergents. [Figure 7]Figure 1 shows ECL signals from an ECL sandwich immunoassay of extracellular vesicles expressing the CD9 surface protein, comparing signals measured using different BDEA-containing read buffers with different surfactants. For each read buffer, the signal measured immediately after addition of the read buffer (T=0) is compared to the signal measured when the assay product was incubated in the read buffer for 15 minutes (T=15) before measuring the signal. [Figure 8] Figure 1 shows ECL signals from an ECL sandwich immunoassay of extracellular vesicles (EVs) expressing the CD9 surface protein, comparing signals measured using different BDEA-containing read buffers containing either TWEEN 20 or PEG(18) tridecyl ether (PEG18TDE) at various concentrations. Four different concentrations of EVs were tested. [Figure 9] Figures 9A-9D show the mean % ECL loss during antibody screening with analytes sRange (Figure 9A), IL-9 (Figure 9B), Kim-1 (Figure 9C), and MIG (Figure 9D) using BDEA or TPA read buffers. [Figure 10] Figures 10A-10C show the % specific and % nonspecific ECL signals in multiplex assays using BDEA or TPA read buffers with the analytes IL-13 (Figure 10A), MDC (Figure 10B), and TNF-β (Figure 10C). [Figure 11] Figure 11A shows the ECL signal change in an EV assay using BDEA or TPA read buffer with varying concentrations of TRITON X-100. Figure 11B shows the titration curve of known concentrations of EVs tested with two different lots of non-TRITON BDEA read buffer. DETAILED DESCRIPTION OF THE INVENTION

[0021] The present disclosure, as well as additional objects, features, and advantages thereof, will be more fully understood from the following detailed description of certain embodiments.

[0022] As described herein, the ECL coreactant tripropylamine (TPA) has certain drawbacks. For example, TPA has low solubility in water, making it difficult to manufacture, and it is volatile with an unpleasant odor. Therefore, the inventors set out to determine whether it would be possible to find a formulation in which TPA was replaced with another ECL coreactant.

[0023] In an embodiment, the invention provides a compound of the formula (HOCH2CH2)2N-CHR 1 -CHR 2 -CHR 3 -R 4 ECL coreactants and formulations comprising alkyldiethanolamines of the formula: [ka] In the formula, R 1 , R 2 , R 3 and R 4 is each independently -H, -CH3, -CH2CH3, or -CH(CH3)2. 1 , R 2 , R 3 are -H and R, respectively. 4 =-CH(BDEA). In an embodiment, R 1 , R 2 , R 3 , and R 4 are each H.

[0024] The present invention includes reagents and formulations containing BDEA or DBAE, preferably BDEA, as an ECL coreactant, providing an alternative to the use of TPA as an ECL coreactant. These coreactants are highly water-soluble, non-volatile, and have little to no odor, thus addressing the major limitations of TPA. The inventors have discovered preferred compositions containing new coreactants that can provide ECL signal generation comparable to TPA-containing compositions while minimizing assay background signal. The compositions of the present invention also offer additional unexpected advantages over conventional TPA-containing compositions, including: (i) the ECL signal is less sensitive to lot-to-lot variations in coreactant components, such as variations in trace levels of contaminants; (ii) the ECL signal is less sensitive to changes in pH, temperature, and salt concentration; (iii) unlike TPA-containing compositions, the ECL signal is insensitive to the presence or absence of detergents and does not require the presence of harsh aromatic detergents such as TRITON X-100 (which can disrupt certain analytes, particularly those containing lipid membranes) for optimal signal generation; and (iv) reduced ECL signal loss, an indicator of a low off-rate between the analyte and the binding reagent. These unexpected advantages, in turn, provide greater robustness than TPA-containing compositions against sources of lot-to-lot variability in manufacturing and run-to-run variability when performing assays. Reducing these sources of assay variability is highly desirable in the field. See, e.g., Lee et al., 2006. In addition, the unexpectedly low sensitivity to the presence of surfactants allows the surfactant composition and concentration to be tailored to achieve specific desired attributes, such as control of the surface tension and meniscus shape of the solution, and the ability to maintain or disrupt the association of components in the assay mixture.

[0025] The present invention includes (i) compositions described herein, (ii) assay reagents comprising one or more of the components of these compositions, (iii) kits containing one of these compositions or reagents and, optionally, other assay components, and (iv) methods of using these compositions, reagents, and / or kits to perform ECL processes or measurements.

[0026] In the discussion and claims herein, the term "about" modifying the amount of a component or reactant of the present invention used refers to variations in the numerical quantity that may occur, for example, through typical measuring and liquid handling procedures used to make concentrates or using solutions, through inadvertent errors in these procedures, through differences in the manufacture, source, or purity of the components used to make the composition or carry out the method, etc. The term "about" also encompasses amounts that differ due to different equilibrium conditions of the composition resulting from a particular initial mixture. Whether modified by the term "about," the claims include equivalents to the amount. In one embodiment, the term "about" means within 10% of the reported numerical value, preferably within 5% of the reported numerical value.

[0027] As used herein, unless otherwise specified, a composition containing a component connected by the term "or" encompasses compositions containing combinations of the components. For example, a composition containing "x," "y," or "z" includes, but is not limited to, compositions containing "x," "y," and "z," compositions containing "x" and "y," compositions containing "x" and "z," and compositions containing "y" and "z."

[0028] As used herein, the terms "substantially" or "substantial" are equally applicable when used in the negative sense to refer to the complete or nearly complete absence of an action, characteristic, property, state, structure, article, or result. For example, a surface that is "substantially" flat will either be completely flat or nearly flat so as to have the same effect as if it were completely flat.

[0029] As used herein, terms such as "a," "an," and "the" are not intended to refer to only a single entity, but include general classes of which specific examples can be used as illustrations.

[0030] Reference herein to any range of values ​​expressly includes each numerical value encompassed by that range, including fractional and integer numbers. For purposes of illustration, reference herein to a range of "at least 50" or "at least about 50" includes integers such as 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, and the like, and decimals such as 50.1, 50.2, 50.3, 50.4, 50.5, 50.6, 50.7, 50.8, 50.9, and the like. For further explanation, reference herein to a range of "less than 50" or "less than about 50" includes integers such as 49, 48, 47, 46, 45, 44, 43, 42, 41, 40, and the like, and decimals such as 49.9, 49.8, 49.7, 49.6, 49.5, 49.4, 49.3, 49.2, 49.1, 49.0, and the like. As a further explanation, references herein to the range "5 to 10" include the integers 5, 6, 7, 8, 9, and 10, as well as the decimals 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, etc.

[0031] Substances that can be induced to release ECL may be referred to as being ECL active or having ECL activity. They may also be referred to as ECL-active species, ECL moieties, ECL labels, ECL-labeled compounds, or ECL-labeled substances. When utilized in certain compositions, reagents, kits, methods, or system embodiments according to the present disclosure, it is within the scope of these ECL-active species to be linked to other molecules, particularly components of biochemical or biological assays, such as analytes or analogs thereof, binding partners of analytes or analogs thereof, additional binding partners of such binding partners, or reactive components capable of binding to analytes, analogs thereof, or binding partners described herein. The ECL-active species described herein may also be linked to a combination of one or more binding partners and / or one or more reactive components. In certain enzyme assays, the ECL-active species may be linked to an enzyme substrate. In the context of excitation of ECL measurement, the ECL-active species may be described as "bound" or "free." In this context, "bound" refers to an ECL-active species held in proximity to the electrode used to induce ECL, e.g., a species immobilized directly on the electrode, held through binding interactions with other species immobilized on the electrode, or present on the surface of beads collected on the surface of the electrode. In contrast, in this context, "free" refers to a label that is free and diffusing in a bulk liquid medium in contact with the electrode, such as a solution, suspension, or emulsion containing the ECL-active species.

[0032] Similarly, the aforementioned "compositions," hereinafter sometimes "ECL compositions," or "systems," containing unstable, metastable, and other intermediate species formed during the ECL reaction, such as the aforementioned excited-state ECL moieties, are within the scope of this disclosure. In addition, while emission of visible light is an advantageous feature of certain embodiments of this disclosure, compositions (hereinafter sometimes "ECL compositions") or systems that emit other types of electromagnetic radiation, such as infrared or ultraviolet radiation, X-rays, microwaves, and the like, are within the scope of this disclosure. The use of the terms "electrochemiluminescence," "electrochemiluminescent," "luminescence," "luminescent," and "luminescence" in the context of this disclosure does not require that the emission be light, but it is recognized that the emission may be such other forms of electromagnetic radiation.

[0033] The compositions of the present disclosure may contain acidic or basic components that can exist in protonated or deprotonated form, for example, depending on the pH of the aqueous composition.For example, a reference to an amine or other base (such as BDEA, DBAE, or Tris) may refer to either the unprotonated (amine) form or the protonated (ammonium) form.Similarly, a reference to a phosphate may refer to either phosphoric acid or the deprotonated form of the phosphate.

[0034] The present disclosure relates to an ECL reading buffer, an assay composition, and a kit containing the same, as well as a method for using the same. A kit refers to a set of components that are provided together to be used, for example, to make a composition, to manufacture a device, or to perform a method. A kit can include one or more components. The components of the kit can be provided in one package or multiple packages, and each package can contain one or more components.

[0035] In embodiments, the compositions of the present disclosure are used in an assay system in place of, in combination with, or as a substitute for a composition containing TPA.

[0036] One aspect of the present disclosure relates to improved ECL assay compositions containing an ECL coreactant, where the ECL coreactant is BDEA or DBAE, preferably BDEA. These ECL assay compositions provide a suitable environment for inducing an ECL label to release ECL and for sensitively measuring the ECL label via ECL measurement. The ECL assay compositions of the present disclosure may optionally contain additional components, including pH buffers, detergents, preservatives, surfactants, antifoaming agents, ECL active species, salts, chelating agents, acids, bases, metal ions and / or metal chelating agents, and / or additional coreactants.

[0037] The ECL assay compositions of the present disclosure may also include biological assay components, including antibodies, antibody fragments, proteins, enzymes, enzyme substrates, enzyme inhibitors, cofactors, antigens, haptens, lipoproteins, liposaccharides, cells, intracellular components, cellular receptors, membrane fragments, exosomes, extracellular vesicles, viruses, nucleic acids, nucleic acid analogs (including protein nucleic acids or analogs with non-natural linkers such as non-natural nucleotide bases), antigens, lipids, glycoproteins, carbohydrates, peptides, amino acids, receptors, hormones, binding reagents, protein-binding ligands, ligands, pharmacological agents, membrane vesicles, liposomes, organelles, bacteria, fungi, or combinations thereof, which may optionally be labeled with an ECL label. These biological assay components may be in non-immobilized form or may be immobilized on a solid surface, including the surface of a solid phase used in solid-phase binding assays, including, but not limited to, the surface of a slide, chip, well, assay cell, tube, or other container, bead, or microparticle. The ECL assay composition can also include the analyte to be measured in the biological assay and / or a sample or component of a sample that includes the analyte to be measured by the biological assay.

[0038] The present disclosure also relates to an ECL read buffer, a composition comprising an ECL coreactant and, optionally, one or more additional components of the ECL assay composition described herein, which is useful for use in an assay for measuring an analyte in a sample. The ECL read buffer is then used by itself or in combination with other assay components to form the ECL assay composition described herein. In one embodiment, the ECL read buffer comprises (i) an ECL coreactant, (ii) a pH buffer component that maintains the pH of the read buffer within a defined range, (iii) an additional ionic component, and, optionally, (iv) a surfactant.

[0039] The ECL assay compositions and ECL read buffers of the present disclosure can be aqueous in nature or substantially aqueous (e.g., >75 wt. %, or preferably >85 wt. % water), although in some applications it may be desirable to add an organic cosolvent such as DMSO, DMF, formamide, ethylene glycol, propylene glycol, methanol, ethanol, glycerol, or other alcohols. In embodiments of the present disclosure, the ECL read buffer (or one or more components thereof) is provided in dry form and is converted by the user into a liquid reagent by adding an appropriate solvent or matrix (preferably water or an aqueous medium), preferably by adding a defined amount of the appropriate solvent or matrix to produce a solution with a pre-specified concentration of the solution components.

[0040] 5.1 ECL coreactants Applicants have discovered ECL read buffers and ECL assay compositions containing non-TPA coreactants that provide comparable or better performance than TPA in some applications.

[0041] In embodiments, the non-TPA co-reactant has the formula (HOCH2CH2)2N-CHR 1 -CHR 2 -CHR 3 -R 4 ) alkyldiethanolamine, [ka] In the formula, R 1 , R 2 , R 3 and R 4 are each independently -H, -CH, -CHCH, or -CH(CH). 1 , R 2 , R 3 are -H and R, respectively. 4 =-CH(BDEA). In an embodiment, R 1 , R 2 , R 3 , and R 4 are each H.

[0042] Non-TPA coreactants include BDEA or DBAE (structures shown below). Preferably, the non-TPA coreactant is BDEA. [ka]

[0043] Compared to TPA, BDEA and DBAE have low volatility (vapor pressure less than 2 mm Hg at room temperature), high boiling points (above 200°C at atmospheric pressure), good water solubility, low odor, and low flammability. Applicant has discovered that as an ECL coreactant, BDEA has the surprising advantage of being relatively unaffected by its concentration. This coreactant can be combined with other components to form compositions, as discussed below. This includes adding other ECL coreactants, such as TPA, or N,N-dibutylethanolamine, or both.

[0044] The concentration of the BDEA or DBAE coreactant in the ECL assay composition and / or read buffer of the present invention can be from about 10 mM to about 800 mM, from about 75 mM to about 400 mM, from about 75 mM to about 300 mM, from about 100 mM to about 300 mM, from about 100 mM to about 225 mM, from about 100 mM to about 150 mM, from about 100 mM to about 175 mM, about 150 mM, or about 125 mM. The concentrations can be selected such that in an ECL assay using the composition and / or read buffer, the specific ECL signal is relatively insensitive to small changes in coreactant concentration, e.g., a concentration change of about 0.5-fold to about 5-fold, or about 0.6-fold to about 2.5-fold, or about 0.7-fold to about 1.2-fold, or about 0.8-fold to about 1.5-fold, or about 0.8-fold to about 1.4-fold, or about 0.8-fold to about 1.2-fold of the nominal value results in a change in the specific ECL signal of about 5% or less, about 10% or less, or about 20% or less.

[0045] In one embodiment, the ECL assay composition and / or read buffer are such that the ECL generation step in an ECL assay using them is relatively insensitive to temperature change, e.g., a temperature change from 18°C ​​to 30°C during the step of generating ECL from an ECL label in the presence of the ECL assay composition results in a change in specific ECL of less than 10%, less than 20%, or less than 30%, or the slope of the change in ECL at temperatures above this range is less than 2% per°C, or less than 10% per°C.

[0046] In embodiments of the disclosed compositions, the ECL coreactant is BDEA. In embodiments, the ECL coreactant is DBAE. In embodiments, the ECL generated by the ECL label in the presence of the composition changes, on average, by less than 1% per degree Celsius over a temperature range of 18°C ​​to 30°C. In embodiments, the ECL is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode.

[0047] In embodiments, the concentration of the ECL coreactant in the composition is about 10 mM to about 800 mM. In embodiments, the concentration of the ECL coreactant is about 75 mM to about 300 mM. In embodiments, the concentration of the ECL coreactant is about 100 mM to about 150 mM. In embodiments, a change in the concentration of the ECL coreactant from 0.8 to 1.2 times its nominal value provides less than a 10% change in ECL generated by the ECL label in the presence of the composition. In embodiments, the ECL is generated in the form of an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode.

[0048] 5.2 pH buffer The ECL assay compositions and read buffers of the present invention containing coreactants are preferably designed to maintain pH within a desired range. To achieve pH control, these compositions and buffers may contain pH buffer components. Materials that can act as pH buffer components to maintain solutions within a specific pH range are well known in the art. Examples of suitable pH buffer components include tris(hydroxymethyl)aminomethane (also referred to herein as "Tris"), phosphate, HEPES, glycylglycine (also referred to as GlyGly), borate, acetate, and citrate. Preferred pH buffer components are Tris and phosphate. While a mixture of Tris and BDEA (or DBAE) provides efficient generation of ECL in the presence of an ECL label, Tris is particularly preferred because it provides a very low ECL background signal in the absence of an ECL label. In one embodiment of the invention, the buffer components used in the ECL assay composition and / or read buffer are selected so that the specific signal of an ECL assay using the composition is greater than, or at least about 20% greater than, or at least about 50% greater than, the signal using the phosphate composition, and the read buffer background signal in the absence of ECL label is less than, or at least about 20% less than, or at least about 50% less than, or at least about 80% less than, the background of a read buffer using the phosphate composition, compared to a similar composition in which the buffer components are replaced with the same concentration of phosphate.

[0049] The concentration of the pH buffering component in the ECL assay composition and / or reading buffer of the present invention can be about 10 mM to about 800 mM, about 50 mM to about 400 mM, about 100 mM to about 300 mM, about 150 mM to 250 mM, or about 200 mM.

[0050] The ECL assay composition or read buffer can be designed or adjusted to have a pH within a defined range. The composition can have a pH ranging from about 6 to about 10, about 6 to about 9, about 7 to about 8, about 7.6 to about 7.9, or about 7.8. In one embodiment, the formulation and pH of the ECL assay composition and / or read buffer are selected so that the ECL signal in an ECL assay using the composition is insensitive to changes in pH, e.g., a change in the specific ECL signal of less than 5% or less than 10% per 0.1 increase or decrease in pH, or a change in the ECL signal of less than 10% per pH unit or less than 25% per pH unit over this pH range.

[0051] In embodiments of the disclosed compositions, the pH buffering component is phosphate, HEPES, glycylglycine, borate, acetate, or citrate.

[0052] In an embodiment of a composition of the present disclosure, the pH buffer component is Tris. In an embodiment, the pH buffer component is Tris and the composition includes a surfactant. In an embodiment, the surfactant is a nonionic surfactant. In an embodiment, the surfactant is a nonionic surfactant containing a phenol ether. In an embodiment, the surfactant is TRITON X-100. In an embodiment, the surfactant is a nonionic surfactant without an aromatic group. In an embodiment, the surfactant is a nonionic surfactant without a phenol ether. In an embodiment, the composition does not disrupt lipid bilayer membranes. In an embodiment, the surfactant is a nonionic surfactant such as KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(n) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the surfactant comprises an alkyl ether-PEG. In embodiments, the surfactant is a linear alkyl ether-PEG. In embodiments, the surfactant is PEG(10) tridecyl ether, PEG(12) tridecyl ether, or PEG(18) tridecyl ether. In embodiments, the surfactant is TWEEN-20. In embodiments, the surfactant is PEG(18) tridecyl ether. "PEG" refers to the polyethylene glycol moiety in the detergent, and PEG(n) refers to a polyethylene glycol moiety containing n monomer subunits.

[0053] In embodiments of the compositions of the present disclosure, the pH of the composition is about 6 to about 9. In embodiments, the pH is about 7 to about 8. In embodiments, the pH is about 7.6 to about 7.9. In embodiments, the pH is about 7.8. In embodiments, the slope of the change in ECL with pH generated by the ECL label in the presence of the composition is less than 10% per pH unit. In embodiments, the ECL is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode.

[0054] In embodiments of the composition, the concentration of the pH buffering component is from about 10 mM to about 800 mM. In embodiments, the concentration of the pH buffering component is from about 100 mM to about 300 mM. In embodiments, the concentration of the pH buffering component is from about 150 mM to about 250 mM.

[0055] In embodiments of the composition, the pH buffering component is not phosphate, and the composition provides at least a 20% increase in ECL generated by the ECL label and / or at least a 20% decrease in background ECL generated in the absence of the ECL label, compared to the same composition containing phosphate as the pH buffering component. In embodiments, the ECL from the ECL label is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode, and / or the ECL in the absence of the ECL label is generated at a carbon-based electrode.

[0056] 5.3 Ions and Salts The compositions and / or read buffers of the present disclosure may contain one or more ions or salts (including or in addition to the pH buffering components described herein) at various concentrations. An important and unexpected discovery in the development of the non-TPA-containing compositions and read buffers of the present invention is the beneficial effect of high ion concentrations, particularly the beneficial effect of high ion concentrations on reducing nonspecific binding of labeled species, compared to ion concentrations typically used in similar TPA-containing compositions. Salts that can be added to increase ion concentrations are well known in the art and include those containing the cation Li. + , Na + , K. + , Rb + , Cs + , Mg +2 , Ca +2 , NH4 + (Preferably Li + , Na + , and K. + ), and / or anion F - , Cl - , Br - , I - , phosphates, sulfates, borates (preferably Cl- Salts that can be used include potassium chloride (KCl), sodium chloride (NaCl), lithium chloride (LiCl), and combinations thereof. In one embodiment of the present invention, the identity and concentration of the ionic species in the non-TPA-containing ECL assay composition and / or read buffer are selected so that the specific signal of the ECL assay is about 50% to about 200%, or about 75 to about 125%, of the signal using the TPA composition, and the nonspecific background signal is about 50% to about 200%, or about 75 to about 125%, of the nonspecific background signal using the TPA composition, compared to a similar ECL assay composition in which the non-TPA coreactant is replaced with TPA.

[0057] ECL assay compositions and / or read buffers of the invention containing coreactants can contain salts, such as those described herein, such that the total concentration of anionic species is about 250 mM or more, about 500 mM or more, about 750 mM or more, about 1000 mM or more, about 250 mM to about 1400 mM, about 500 mM to about 1200 mM, or about 1050 mM. Such compositions and / or read buffers can also contain chloride ions (Cl). - ) is about 250 mM or more, about 500 mM or more, about 750 mM or more, about 1000 mM or more, about 250 mM to about 1400 mM, about 500 mM to about 1200 mM, or about 1050 mM.

[0058] Cl -In embodiments containing components, the disclosed compositions may include NaCl, KCl, LiCl, or a mixture thereof. In this embodiment, the concentration of NaCl may be about 200 mM to about 1400 mM, about 600 mM to about 1200 mM, or about 800 mM; the concentration of KCl may be about 50 mM, about 200 mM to about 1400 mM, about 500 mM, about 600 mM to about 1200 mM, or about 800 mM; and the concentration of LiCl may be about 200 mM to about 1400 mM, about 600 mM to about 1200 mM, or about 800 mM. The disclosed compositions may include NaCl at a concentration of about 800 mM, and the concentration of KCl may be about 50 mM. The disclosed compositions may also include LiCl at a concentration of about 800 mM, and the concentration of KCl may be about 50 mM.

[0059] In one embodiment of the disclosed composition, the composition may have an ionic strength of about 0.3M to about 1.9M or about 1.4M, about 0.5M to about 1.2M or about 1.0M, about 0.8M to about 1.2M, about 1.0M to 1.2M, or about 1.1M.

[0060] In an embodiment of the composition of the present disclosure, the ionic component comprises chloride ions. In an embodiment, the ionic component comprises NaCl, KCl, LiCl, or a combination of any two or any three of their salts. In an embodiment, the ionic component comprises NaCl. In an embodiment, the ionic component comprises KCl.

[0061] In embodiments, the composition has an ionic strength greater than about 0.3 M. In embodiments, the composition has an ionic strength greater than about 0.5 M. In embodiments, the composition has an ionic strength greater than about 0.8 M. In embodiments, the composition has an ionic strength greater than about 1.0 M.

[0062] In embodiments, the composition comprises chloride ions, and the concentration of chloride ions is greater than about 0.25 M. In embodiments, the composition comprises chloride ions, and the concentration of chloride ions is greater than about 0.5 M. In embodiments, the composition comprises chloride ions, and the concentration of chloride ions is greater than about 0.75 M. In embodiments, the composition comprises chloride ions, and the concentration of chloride ions is greater than about 1.0 M.

[0063] In embodiments, non-specific binding (NSB) in immunoassays using compositions containing ionic components is lower compared to the same compositions without the ionic components.

[0064] 5.4 Surfactants An unexpected aspect of BDEA- and DBAE-based ECL compositions and read buffers is the insensitivity of ECL generation to the presence, concentration, or structure of surfactants. In contrast, TPA-based compositions require the presence of surfactants for optimal signal generation. In particular, TPA provides optimal ECL generation in the presence of surfactants containing aromatic moieties, such as the phenol ether moiety of TRITON X-100.

[0065] Thus, the coreactant-containing ECL assay compositions and / or read buffers of the present invention include compositions that (i) are detergent-free or (ii) contain a detergent, but only at a concentration below the detergent's critical micelle concentration (cmc). The critical micelle concentration (cmc) is known as the concentration of a detergent above which micelles form and any additional detergent added to the composition goes to the micelles. The coreactant-containing ECL assay compositions and / or read buffers of the present invention also include (i) compositions that are free of aromatic surfactants, or more specifically surfactants containing a phenol ether moiety, or more specifically TRITON X-100, or (ii) compositions that contain a surfactant described in (i), but only at a concentration below the detergent's critical micelle concentration (cmc).

[0066] Alternatively, including a surfactant, for example, to provide lower surface tension or to break down undesirable molecular aggregates, may still be advantageous in some applications. Possible surfactants include aromatic surfactants such as TRITON X-100 and / or non-aromatic surfactants. Possible surfactants include nonionic and ionic surfactants. Possible nonionic surfactants include those known under the trade names NONIDET, BRIJ, TRITON, TWEEN, THESIT, LUBROL, GENAPOL, PLURONIC, TETRONIC, F108, and SPAN. Preferably, the surfactant is included at a concentration that provides a solution with an air-liquid surface tension of 50 dynes / cm or less, about 40 dynes / cm or less, or about 35 dynes / cm or less. Preferably, the included surfactant is present at a concentration equal to or greater than their cmc, about two times their cmc, or about five times their cmc. In embodiments, the compositions and / or read buffers of the present disclosure contain a surfactant at a concentration of about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% by weight per volume of the composition. In embodiments, the compositions and / or read buffers of the present disclosure contain a surfactant at a concentration of about 0.1 mM to about 10 mM, about 0.2 mM to about 8 mM, about 0.3 mM to about 5 mM, about 0.4 mM to about 0.8 mM, about 0.5 mM to about 0.6 mM, about 1 mM, or about 0.5 mM. In embodiments, the detergent contained in the ECL coreactant-containing composition and / or read buffer is present at a concentration of about 0.1 mM, about 0.2 mM, about 0.3 mM, about 0.4 mM, about 0.5 mM, about 0.6 mM, about 0.7 mM, about 0.8 mM, about 0.9 mM, about 1 mM, about 2 mM, about 3 mM, about 4 mM, about 5 mM, about 6 mM, about 7 mM, about 8 mM, about 9 mM, or about 10 mM. In embodiments, the detergent concentration is about 0.2 mM to about 10 mM. In embodiments, the detergent concentration is about 0.5 mM to about 8 mM.In embodiments, the surfactant concentration is about 1.0 mM to about 5 mM. In embodiments, the surfactant concentration is about 0.5 mM. In embodiments, the surfactant concentration is about 1.0 mM. In embodiments, the surfactant concentration is 5.0 mM. In embodiments, the surfactant concentration is about 10 mM. In embodiments, the surfactant concentration is above the critical micelle concentration (cmc) of the surfactant.

[0067] Advantageously, surfactants can be selected for inclusion in compositions and / or read buffers that offer some desirable attributes of surfactants, such as low surface tension (and better-controlled meniscus angles), but are less denaturing than TRITON X-100 with respect to a particular biological assay target. The present invention includes embodiments in which mild surfactants are included that do not disrupt analytes containing lipid bilayer membranes. Such mild surfactants include, but are not limited to, (i) oligomers of ethylene glycol and / or propylene glycol, or (ii) alkyl chains linked to oligomers of ethylene glycol and / or propylene glycol via ether groups. Such mild surfactants may also be from the classes of surfactants known under the trade names BRIJ, TWEEN, PLURONIC, or KOLLIPHOR. Such mild surfactants also include KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-5-decyne-4,7-diol ethoxylate, HO(CH2CH2) 18 C 13 H 27 (also known as PEG(18) tridecyl ether), BRIJ L4, BRIJ 58, or TWEEN 20. Such mild surfactants may also be specific surfactants such as KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether. Preferably, the surfactant does not contain an ester linkage.

[0068] In embodiments, the composition of the present disclosure includes a nonionic surfactant that does not contain phenol ether. In embodiments, the composition does not disrupt lipid bilayer membranes. In embodiments, the surfactant is a nonionic surfactant such as KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the surfactant is TWEEN 20. In embodiments, the surfactant is PEG(18) tridecyl ether. In embodiments, the pH buffering component is phosphate, HEPES, glycylglycine, borate, acetate, or citrate.

[0069] 5.5 ECL Labels and Other Assay Reagents The compositions of the invention may comprise a variety of assay reagents and / or components useful for performing assay measurements, which may optionally be labeled with an ECL label, including whole cells, cell surface antigens, intracellular particles (e.g., organelles, or membrane fragments), exosomes, extracellular vesicles, liposomes, membrane vesicles, viruses, prions, house dust mites or fragments thereof, viroids, antibodies, antigens, haptens, fatty acids, nucleic acids (and synthetic analogs), proteins (and synthetic analogs), lipoproteins, polysaccharides, inhibitors, cofactors, haptens, cellular receptors, receptor ligands, lipopolysaccharides, glycoproteins, peptides, polypeptides, enzymes, enzyme substrates, enzyme products, second messengers, cellular metabolic products, hormones, pharmacological agents, synthetic organic molecules, organometallic molecules, sedatives, barbiturates, alkaloids, steroids, vitamins, amino acids, sugars, lectins, recombinant or derived proteins, biotin, avidin, and / or streptavidin. Assay reagents may be useful, for example, as binding reagents or enzyme substrates in binding or enzyme assays.

[0070] The composition of the present invention may contain an ECL label. Useful ECL labels include those that generate ECL in the presence of TPA as a coreactant. Examples of ECL labels that can be used include luminescent organometallic compounds, such as those containing Ru, Os, Ir, Re, or lanthanide metals (see, for example, the labels described in U.S. Pat. Nos. 5,714,089, 6,316,607, 6,808,939, and 9,416,150). Preferred ECL labels are ruthenium- or osmium-containing organometallic species. These ruthenium- or osmium-containing organometallics may contain ruthenium or osmium chelated to polypyridyl ligands (most preferably bipyridine, phenanthroline, and / or their substituted derivatives). Most preferably, the ECL label comprises ruthenium-tris-bipyridine, where the bipyridine ligand is optionally substituted, for example, with a linking group for coupling the label to an assay reagent and / or with a hydrophilic substituent to minimize nonspecific binding. In one embodiment, the ECL label used is a ruthenium tri-bipyridine derivative containing one or two 4,4'-bis(sulfomethyl)-bipyridine ligands (such as those described in U.S. Pat. No. 6,808,939). The ECL label can be linked to the assay reagents described herein.

[0071] In certain embodiments in which the compositions of the present invention are used in analyte binding assays, the label can be linked to a binding component of the assay, such as the analyte or an analog thereof, a binding partner of the analyte or an analog thereof, an additional binding partner of such a binding partner, or a reactive component capable of binding to the analyte, its analog, or a binding partner described herein. The labels described herein can also be linked to a combination of one or more binding partners and / or one or more reactive components. The reactive component can be used to indirectly immobilize or label another assay component. For example, in a binding assay using a binding reagent for the analyte, the binding reagent can be directly immobilized or labeled, or can be indirectly labeled or immobilized using a reactive component. Such approaches are well known in the art and include (i) the use of a secondary binding reagent, such as an anti-species antibody (e.g., an anti-analyte antibody), to label or immobilize the assay reagent, or (ii) the use of labeled or immobilized streptavidin to label or immobilize a biotin-binding reagent conjugate.

[0072] 5.6 Other Components The ECL assay compositions and / or read buffers of the present disclosure may contain one or more other components at various concentrations. In compositions consisting essentially of the listed components, such compositions contain the listed components and components that do not physically affect the basic and novel characteristics of the composition. Components that do not physically affect the basic and novel characteristics of the composition are those that do not alter the essential ECL characteristics of the composition (e.g., generation of an ECL signal). Non-limiting examples of components that may be included in the composition include cosolvents (such as those described herein), preservatives and / or biocides (e.g., azides, 5-chloro-2-methyl-4-isothiazolin-3-one, and / or 2-methyl-isothiazolin-3-one), antifoaming agents (silicone and / or non-silicone-based agents, such as Antifoams 204, A, B, C, Y-30, and / or SE-15 from Millipore Sigma), colorants, and tracer chemicals added to provide a chemical fingerprint to verify the origin or authenticity of the reagent. In embodiments, perchlorate compounds are included.

[0073] 5.7 Concentrated and dried forms Another embodiment of the present disclosure relates to a dried or concentrated reagent composition that can be diluted with a diluent (e.g., water or an aqueous solution) to form the ECL assay composition and / or read buffer described above. By way of example, such a concentrated reagent composition may have reagent components in dry or liquid form at a concentration higher than the concentration of the reagent in the desired ECL assay composition and / or read buffer (e.g., the concentration may be 2X or more, 4X or more, or 10X or more of the desired concentration). The present invention also includes a method of forming the ECL assay composition and / or read buffer of the present invention, comprising mixing a dried or concentrated composition with a diluent (an aqueous solution, preferably water). Preferably, a predefined amount of the dried or concentrated composition is provided, and a predefined amount of the diluent is added. In embodiments in which a concentrated composition is used, the ratio of the volume of the diluent to the volume of the concentrated composition may be about 1 or more, about 2 or more, or about 3 or more.

[0074] The formulation of the disclosed ECL assay composition and / or read buffer can be selected to be relatively insensitive to component concentration or dilution, for example, if the user does not accurately measure the amount of diluent used to rehydrate dried reagents or dilute a concentrated composition. In one embodiment, the composition and / or read buffer is configured so that the specific ECL signal from an ECL assay remains relatively unchanged, or changes by less than 10%, or by less than 20%, or by less than 30%, over a concentration range of 0.8-1.2 times the nominal concentration of the component.

[0075] In an embodiment of the present disclosure, the composition comprises a liquid diluent, hi an embodiment, the liquid diluent is water and the composition is substantially aqueous.

[0076] In an embodiment of the present disclosure, the composition does not include a liquid diluent and the composition is provided in a dry form.

[0077] 5.8 Kits One aspect of the present disclosure relates to a kit comprising one or more components of the disclosed ECL assay composition and / or read buffer in one or more containers. Preferably, at least one of the containers contains BDEA (or, alternatively, DBAE). Suitable containers that can be used include, but are not limited to, glass and / or plastic containers and plastic and / or foil pouches. The kit packaging or component containers may bear information about the contents of the kit or container, or instructions for proper storage and / or use in an assay. These components can optionally be combined with additional reagents to form the disclosed ECL assay composition and / or read buffer. The kit may also include additional assay-related components, such as ECL labels, ECL-labeled assay reagents, enzymes, binding reagents, electrodes, assay plates, etc. The kit may include components in liquid or dry form.

[0078] Another aspect of the present disclosure relates to kits containing two or more components that, when mixed, form the ECL assay composition or read buffer described herein. The components can be solid and / or liquid, and preferably, at least one component is liquid. The present invention also includes methods of forming an ECL assay composition and / or read buffer, comprising mixing the components in such kits. In one embodiment, predetermined amounts of each component are mixed. Such predetermined amounts of components may be provided pre-measured in separate containers. In one embodiment, the method also includes providing one or more additional components not part of the kit and combining them with the kit components. In one example, the additional component is a liquid diluent (e.g., water).

[0079] Another aspect of the present disclosure relates to a kit for performing an assay, comprising an ECL read buffer and one or more additional assay components in one or more containers. In such a kit, the ECL read buffer can be provided as a single component or as two or more of the above components. Additional assay components used to perform the assay that may be part of the kit include, but are not limited to, (i) an assay instrument, (ii) assay consumables, (iii) additional assay reagents, and (iv) assay samples for use, for example, as calibration standards or assay controls.

[0080] Examples of assay consumables that may be included in the kit are assay modules designed to house samples and / or reagents during one or more steps of the assay, pipette tips and other consumables for transferring liquid samples and reagents, covers and seals for the assay modules and other consumables used in the assay, racks for holding other assay consumables, labels (including human-readable or machine-readable formats such as barcodes, RFID, etc.) for identifying samples or other assay consumables, and media (including paper and electronic media) for providing information about the assay and / or instructions for performing the assay.

[0081] Assay modules can include tubes, cuvettes, wells, multiwell plates, cartridges, lateral flow devices, flow cells, etc. The kit can include an assay module having one or more assay electrodes, such as an assay plate or multiwell assay plate according to U.S. Pat. Nos. 7,842,246 and 6,977,722, entitled "Assay Plates, Reader Systems and Methods for Luminescence Test Measurements," or an assay cartridge according to U.S. Pat. No. 9,731,297, entitled "Assay Cartridges and Methods of Using the Same," each of which is incorporated herein by reference. Preferably, the assay electrodes include carbon electrodes, which may be carbon ink electrodes. According to one embodiment, the kit includes an assay module, which serves as a container for one or more other assay components provided in the kit. In such embodiments, the one or more other assay components may include an ECL read buffer or one or more components of an ECL read buffer, for example, a read buffer or a component thereof may be provided in one or more of the multiwell plate wells, optionally in dried form.

[0082] According to another embodiment, the kit includes an assay module, which serves as a container for at least one binding reagent. Optionally, the binding reagent is immobilized on the assay module. Optionally, a plurality of binding reagents are immobilized as an array of binding reagents within the assay module. The immobilized binding reagent and / or the array of immobilized binding reagents may be immobilized on an electrode (which may be a carbon electrode or a carbon ink electrode) within the assay module.

[0083] According to another embodiment, the kit includes a multi-well assay plate having a plurality of wells, the assay plate being used as a container for at least one binding reagent. Optionally, the binding reagent is immobilized on the plate. A plurality of wells in the plate may have immobilized binding reagents therein. The binding reagent in each of these wells may be the same as in all, some, or none of these wells. Optionally, the plurality of binding reagents are immobilized on the wells as an array of respective binding reagents. The immobilized binding reagent and / or the array of immobilized binding reagents may be immobilized on an electrode (which may be a carbon-based electrode, or more specifically, a carbon ink electrode) in the well.

[0084] Binding reagents that may be included in the kit (and / or in the assay module, immobilized or non-immobilized) include, but are not limited to, whole cells, cell surface antigens, intracellular particles (e.g., organelles, or membrane fragments), exosomes, extracellular vesicles, membrane vesicles, liposomes, viruses, prions, house dust mites or fragments thereof, viroids, antibodies, antigens, haptens, fatty acids, nucleic acids (and synthetic analogs), proteins (and synthetic analogs), lipoproteins, polysaccharides, inhibitors, cofactors, haptens, cellular receptors, receptor ligands, lipopolysaccharides, glycoproteins, peptides, polypeptides, enzymes, enzyme substrates, enzyme products, second messengers, cellular metabolic products, hormones, pharmacological agents, synthetic organic molecules, organometallic molecules, sedatives, barbiturates, alkaloids, steroids, vitamins, amino acids, sugars, lectins, recombinant or derived proteins, biotin, avidin, and / or streptavidin.

[0085] In one embodiment, the kit includes a binding reagent comprising a lipid bilayer membrane (provided in the assay module and optionally immobilized therein) and an ECL read buffer (provided in one or more portions), in which the lipid bilayer membrane is not disrupted. The read buffer may be detergent-free. Alternatively, the read buffer may contain a detergent, but at a concentration below the cmc of the detergent. Alternatively, the read buffer may contain a mild detergent (which may be present at a concentration above the cmc) that does not disrupt the lipid bilayer membrane. In embodiments, the lipid bilayer is derived from at least one of an intact cell, a cell lysate, a cell fragment, a cell membrane, a membrane ghost, an organelle, an organelle fragment, an organelle membrane, a virion, a virion fragment, a virion membrane, a liposome, a plasma membrane fragment, an endosome, a clathrin-coated vesicle, an endoplasmic reticulum fragment, a synaptic vesicle, a Golgi apparatus fragment, a membrane subdomain, a mitochondria, a peroxisome, a lysosome, a liposome, an exosome, an extracellular vesicle, a virus particle, a virus-derived membrane-enclosed particle shed from a cell, or an intact lipid membrane body from an organism.

[0086] According to another embodiment, the kit comprises two or more, four or more, eight or more, fifteen or more, or twenty-five or more assay modules or plates. According to one embodiment, the kit is contained in a resealable bag or container (e.g., a container with a zip-lock opening).

[0087] In embodiments, the present disclosure provides a kit comprising the following materials in one or more containers: N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both; a pH buffering component; and an ionic component.

[0088] In embodiments, one or more of the ingredients are provided in dry form. In embodiments, the kit further comprises a liquid diluent. In embodiments, the kit further comprises a surfactant. In embodiments, the kit does not include a surfactant or a liquid diluent.

[0089] In embodiments, the present disclosure provides kits comprising a composition described herein and an assay device, an assay consumable, additional assay reagents, an assay sample, or a combination thereof. In embodiments, the kit comprises an assay device, wherein the assay device is configured for performing an ECL assay. In embodiments, the kit comprises assay consumables, and further comprises an electrode configured for use in an ECL assay. In embodiments, the electrode is a carbon-based electrode. In embodiments, the electrode is a screen-printed carbon ink electrode.

[0090] In embodiments, the kit includes an additional assay reagent, wherein the additional assay reagent is a binding reagent. In embodiments, the binding reagent is labeled with an ECL label. In embodiments, the label is an organometallic ruthenium complex. In embodiments, the kit includes an additional assay reagent, wherein the additional assay reagent is a binding reagent, and wherein the additional assay reagent is immobilized on an electrode. In embodiments, the kit includes an additional assay reagent, wherein the additional assay reagent is a binding reagent, and wherein the additional assay reagent is immobilized as an array on an electrode. In embodiments, the kit includes an additional assay reagent, wherein the additional assay reagent is a binding reagent, and wherein the additional assay reagent is immobilized on particles. In embodiments, the particles are magnetically collectable.

[0091] In embodiments, the kit comprises at least one assay sample, wherein the at least one assay sample comprises an assay calibration sample and / or an assay control sample.

[0092] 5.9 Method Another aspect of the present disclosure relates to methods of using the buffers, reagents, and / or compositions of the present disclosure.

[0093] One embodiment of the present disclosure relates to a method for performing an electrochemiluminescence assay, wherein electrochemiluminescence is induced in the presence of an ECL assay composition and / or a read buffer of the present disclosure. Electrochemiluminescence can be induced using a carbon-based electrode. The method according to this embodiment includes contacting the electrode with the ECL assay composition and / or a read buffer, applying a voltage to the electrode, and measuring the resulting electrochemiluminescence.

[0094] Another embodiment of the present disclosure relates to a method for measuring the amount of ECL label, wherein the label is induced to emit electrochemiluminescence in the presence of an ECL assay composition and / or a read buffer of the present disclosure, and the electrochemiluminescence is measured to determine the amount of ECL label. Electrochemiluminescence can be induced using a carbon-based electrode. The method according to this embodiment includes contacting the electrode with the ECL assay composition and / or read buffer, applying a voltage to the electrode, and measuring the resulting electrochemiluminescence, wherein the amount of ECL label is immobilized or retained in proximity to the electrode. Alternatively, the amount of ECL label can be a component of the ECL assay composition and / or read buffer.

[0095] Another embodiment of the present disclosure relates to a method for measuring the amount of a binding complex containing a binding assay reagent labeled with an ECL label, wherein the label is induced to emit electrochemiluminescence in the presence of an ECL assay composition and / or a read buffer of the present disclosure, and the electrochemiluminescence is measured to measure the amount of the binding complex. The electrochemiluminescence can be induced using a carbon-based electrode. The method according to this embodiment includes: (i) contacting the electrode with a binding reaction solution, the electrode having an immobilized binding reagent immobilized thereon, the binding reaction solution including a labeled binding reagent labeled with an ECL label; (ii) forming a binding complex on the electrode containing the immobilized binding reagent and the labeled binding reagent; (iii) contacting the binding complex on the electrode with an ECL assay composition containing a coreactant; (iv) applying a voltage to the electrode in the presence of the ECL assay composition; and (v) measuring the resulting ECL to measure the amount of the binding complex. Another method according to this embodiment includes: (i) contacting particles (or other solid-phase binding reaction support) with a binding reaction solution, where the particles have immobilized binding reagents immobilized thereon and the binding reaction solution includes a labeled binding reagent labeled with an ECL label; (ii) forming a binding complex on the particles including the immobilized binding reagent and the labeled binding reagent; (iii) contacting the binding complex on the particles with an ECL assay composition containing a coreactant; (iv) collecting the particles on an electrode (e.g., by gravity, centrifugation, filtration, or, in the case of magnetic particles, by use of a magnet); (v) applying a voltage to the electrode in the presence of the ECL assay composition; and (vi) measuring the resulting ECL to determine the amount of binding complex. In a further embodiment, the electrode comprises or consists essentially of platinum.

[0096] The method may use a "wash" assay format, in which step (iii) of the method described herein further comprises washing the electrode (or particles) to remove unbound labeled binding reagent and contacting the electrode (or particles) with an ECL coreactant-containing ECL read buffer to provide an ECL assay composition. Alternatively, a "no-wash" format may be used, in which step (iii) does not include a wash step and comprises combining the binding reaction solution with an ECL coreactant-containing ECL read buffer to form the ECL assay composition. In the alternative "no-wash" format, the binding reaction solution is a coreactant-containing ECL assay composition.

[0097] Another embodiment of the present disclosure includes a method for performing an assay for an analyte using the method for measuring binding complexes described herein. In this embodiment, step (i) of the method may further include (a) contacting the electrode (or particles) with a sample containing an amount of analyte, and / or (b) combining a labeled binding reagent with a sample containing an amount of analyte to form a binding reaction solution. Furthermore, step (v) may include determining the amount of analyte from the amount of binding complex. Binding assay formats that can be performed include, but are not limited to, (i) direct binding assays, in which the labeled binding reagent is the analyte of interest and the immobilized binding reagent is a binding partner of the analyte, and a complex is formed by direct binding of the two reagents; (ii) sandwich binding assays, in which the immobilized and labeled binding reagents are both binding partners of the analyte of interest, and the analyte binds to the two binding partners to form a complex; and (iii) competitive binding assays, in which the immobilized binding reagent is a binding partner of the analyte and the labeled binding reagent is a competitor (e.g., the analyte or an analog of the analyte) that competes with the immobilized binding reagent for binding to the analyte, or the labeled binding reagent is a binding partner of the analyte and the immobilized binding reagent is a competitor that competes with the immobilized binding reagent for binding to the analyte (in competitive formats, the labeled binding complex—formed by direct binding of the immobilized and labeled binding reagent—decreases in amount as the amount of analyte increases). Where immobilized and / or labeled binding reagents are described herein as being binding partners or competitors of an analyte, it is well understood in the art that they can in turn be replaced with reactive components that bind to the analyte's binding partner or competitor, either directly or through an additional reactive component.

[0098] Another embodiment of the present disclosure relates to a method for measuring the amount or activity of an analyte, in which the analyte reacts with, forms a complex with, or competes in a specific binding interaction with a label containing an ECL label, and the label is induced to emit electrochemiluminescence in the presence of an ECL coreactant-containing ECL assay composition and / or a read buffer of the present disclosure, and the electrochemiluminescence is measured to measure the amount or activity of the analyte. Electrochemiluminescence can be induced using a carbon-based electrode. The presence or activity of the analyte causes the label to bind to or be released from the electrode (e.g., via the formation of a specific binding complex or via the cleavage or formation of a chemical bond). A method according to this embodiment can include contacting the electrode with the ECL assay composition and / or read buffer, applying a voltage to the electrode, and measuring the resulting electrochemiluminescence, wherein the amount of ECL label is immobilized or retained in proximity to the electrode. Advantageously, the waveform used to induce ECL in the presence of the ECL assay compositions and / or read buffers of the present disclosure may be short in duration, e.g., the waveform may have a duration of less than 12 seconds, less than 6 seconds, less than 3 seconds, less than 2 seconds, or less than 1 second.

[0099] Approaches for determining the amount and / or concentration of an ECL label or analyte using the amount of ECL measured in an ECL assay are established in the art and may include, for example, using calibration standards and / or calibration curves to establish a relationship between the ECL signal and the amount and / or concentration of the label and / or analyte. Calibration can be performed at different times, for example, during method development, during qualification of a particular lot of assay material, or at the time of assay measurement. Calibration can also be performed using calculations based on the known physical and chemical behavior of the assay components and equipment.

[0100] Embodiments of the present disclosure can be used to test a variety of samples that may contain an analyte or activity of interest. Such samples can be in the form of a solid, emulsion, suspension, liquid, or gas. They can include, but are not limited to, for example, cells (live or dead) and cell-derived products, immortalized cells, cell fragments, cell fractions, cell lysates, organelles, cell membranes, hybridomas, cell culture supernatants (including supernatants from antibody-producing organisms such as hybridomas), wastewater, or drinking water, food, beverages, pharmaceutical compositions, blood, serum, plasma, hair, sweat, urine, feces, tissue, biopsies, waste fluids, separated and / or fractionated samples, separated and / or fractionated liquids, organs, saliva, animal parts, animal by-products, plants, plant parts, plant by-products, soil, minerals, mineral deposits, water, waterers, water sources, residues (gas and liquid) filtered from fluids, swipes, absorbent materials, gels, cytoskeleton, protein complexes, unseparated samples, and the like. The sample may contain or be derived from a fractionated sample, unfractionated cell lysate, endocrine factors, paracrine factors, autocrine factors, cytokines, hormones, cell signaling factors and / or components, second messenger signaling factors and / or components, cell nuclei / nuclei, nuclear fractions, chemicals, chemical compositions, structural biological components, skeletal (ligament, tendon) components, isolated and / or fractionated skeletal components, hair, fur, feathers, hair fractions and / or isolates, skin, skin samples, skin fractions, dermis, endothelium, eukaryotic cells, prokaryotic cells, fungi, yeast, antibodies, antibody fragments, immune factors, immune cells, drugs, therapeutic agents, oils, extracts, mucous membranes, fur, oils, sewage, environmental samples, organic solvents, or air. The sample may further comprise, for example, water, organic solvents (e.g., acetonitrile, dimethyl sulfoxide, dimethylformamide, n-methylpyrrolidone, or alcohol), or mixtures thereof.

[0101] Analytes that may be measured include, but are not limited to, whole cells, cell surface antigens, intracellular particles (e.g., organelles, or membrane fragments), exosomes, extracellular vesicles, liposomes, membrane vesicles, viruses, prions, house dust mites or fragments thereof, viroids, antibodies, antigens, haptens, fatty acids, nucleic acids (and synthetic analogs), proteins (and synthetic analogs), lipoproteins, polysaccharides, inhibitors, cofactors, haptens, cellular receptors, receptor ligands, lipopolysaccharides, glycoproteins, peptides, polypeptides, enzymes, enzyme substrates, enzyme products, second messengers, cellular metabolic products, hormones, pharmacological agents, synthetic organic molecules, organometallic molecules, sedatives, barbiturates, alkaloids, steroids, vitamins, amino acids, sugars, lectins, recombinant or derived proteins, biotin, avidin, streptavidin, or inorganic molecules present in the sample. Activities that may be measured include, but are not limited to, activities of phosphorylases, phosphatases, esterases, transglutaminases, nucleic acid damage activity, transferases, oxidases, reductases, dehydrogenases, glycosidases, ribosomes, protein processing enzymes (e.g., proteases, kinases, protein phosphatases, ubiquitin-protein ligases, etc.), nucleic acid processing enzymes (e.g., polymerases, nucleases, integrases, ligases, helicases, telomerases, etc.), cell receptor activation, second messenger system activation, and the like.

[0102] Whole cells can be animal, plant, or bacterial, and can be viable or dead. Examples include fungi and plant pathogens such as nematodes. The term "intracellular particle" is meant to encompass, for example, intracellular organelles, membrane particles from disrupted cells, cell wall fragments, ribosomes, multienzyme complexes, and other particles that may be derived from an organism. Nucleic acids include, for example, chromosomal DNA, plasmid DNA, viral DNA, and recombinant DNA from multiple sources. Nucleic acids also include RNA, such as messenger RNA, ribosomal RNA, and transfer RNA. Polypeptides include structural proteins, such as enzymes, transport proteins, receptor proteins, and viral coat proteins. Preferred polypeptides are enzymes and antibodies. Particularly preferred polypeptides are monoclonal antibodies. Hormones include, for example, insulin and T4 thyroid hormone. Pharmacological agents include, for example, cardiac glycosides. It is within the scope of this disclosure to include synthetic polypeptides, synthetic nucleic acids, and synthetic materials that chemically resemble biological materials, such as synthetic membranes, vesicles, and liposomes. The foregoing is not intended to be an exhaustive list of biological materials suitable for use in the present disclosure, but is meant only to illustrate the broad scope of the present disclosure.

[0103] In embodiments, the present disclosure provides a method for producing a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In method embodiments, one or more of the ingredients are provided in dry form.

[0104] In embodiments, the present disclosure provides methods for generating ECL comprising contacting an electrode with a composition described herein and an ECL label, applying a voltage to the electrode, and generating ECL.

[0105] In embodiments, the present disclosure provides a method for measuring the amount of an ECL label, comprising contacting an electrode with a composition described herein and an ECL label, applying a voltage to the electrode, generating ECL, measuring the ECL, and determining the amount of label from the measured ECL.

[0106] In embodiments, the present disclosure provides a method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, the method comprising contacting a binding reagent immobilized on an electrode with a labeled binding reagent comprising an ECL label, forming a binding complex on the electrode comprising the immobilized binding reagent and the labeled binding reagent, contacting the binding complex on the electrode with a composition described herein, applying a voltage to the electrode in the presence of the composition, generating ECL, and measuring the ECL to determine the amount of binding complex on the electrode.

[0107] In embodiments, the present disclosure provides a method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, the method comprising: contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label, forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent, contacting the binding complex on the particle with a composition described herein, collecting the particles on an electrode, applying a voltage to the electrode in the presence of the composition, generating ECL, and measuring the ECL to determine the amount of binding complex on the electrode. In embodiments, the method further comprises washing the electrode after formation of the binding complex and before and / or during contacting the electrode with the composition.

[0108] In embodiments, the present disclosure provides a method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, the method comprising: contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label; forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent; contacting the binding complex on the particle with a composition described herein; collecting the particles on an electrode; applying a voltage to the electrode in the presence of the composition; generating ECL; and measuring the ECL to determine the amount of binding complex on the electrode. In embodiments, the particles are magnetically collectable, and the particles are collected on the electrode using a magnetic field. In embodiments, the method further comprises washing the particles after forming the binding complex and before and / or during contacting the particles with the composition.

[0109] In method embodiments, the immobilized binding reagent and the labeled binding reagent bind directly to each other. In method embodiments, the immobilized binding reagent and the labeled binding reagent bind indirectly to each other through other binding species. In embodiments, the immobilized binding reagent and the labeled binding reagent comprise (i) a binding partner of the analyte of interest, (ii) the analyte of interest, or an analog and / or competitor thereof, or (iii) a reactive component capable of binding to species (i) or (ii).

[0110] In embodiments, the present disclosure provides a method for measuring the amount of an analyte, the method comprising: contacting a binding reagent immobilized on an electrode with a labeled binding reagent comprising an ECL label and an analyte (or a sample comprising the analyte); forming a binding complex on the electrode comprising the immobilized binding reagent and the labeled binding reagent; contacting the binding complex on the electrode with a composition described herein; applying a voltage to the electrode in the presence of the composition; generating ECL; and measuring the ECL to determine the amount of the analyte. In embodiments, the method further comprises washing the electrode after formation of the binding complex and before and / or during contacting the electrode with the composition. In embodiments, the method is configured for performing multiplex measurements of multiple analytes, wherein the electrode has an array of immobilized binding reagents for the multiple analytes immobilized thereon.

[0111] In embodiments, the present disclosure provides a method for measuring the amount of an analyte, the method comprising: contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label and the analyte (or a sample comprising the analyte); forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent; contacting the binding complex on the particle with a composition described herein; collecting the particles on an electrode; applying a voltage to the electrode in the presence of the composition; generating ECL; and measuring the ECL to determine the amount of the analyte. In embodiments, the particles are magnetically collectable particles, and the particles are collected on the electrode using a magnetic field. In embodiments, the method further comprises washing the particles after forming the binding complex and before and / or during contacting the particles with the composition. In embodiments, the immobilized binding reagent and the labeled binding reagent comprise (i) a binding partner of the analyte, (ii) the analyte, or an analog and / or competitor thereof, or (iii) a reactive moiety capable of binding to species (i) or (ii).

[0112] In embodiments of the method, the electrode is a carbon-based electrode. In embodiments, the electrode is a screen-printed carbon ink electrode. In embodiments, the ECL is imaged using a camera. In embodiments, the ECL is measured using a photodiode. In embodiments, the electrode is in a multi-well plate assay consumable. In embodiments, the electrode is in a flow cell. In embodiments, the electrode is platinum.

[0113] 5.10 Composition Below are some non-limiting examples of embodiments of the disclosed ECL assay compositions and / or read buffers.

[0114] In one embodiment of the disclosed composition, the composition contains the following components: BDEA at a concentration of about 75 mM to about 300 mM; tris(hydroxymethyl)aminomethane (Tris) buffer at a concentration of zero or about 100 mM to about 300 mM; TRITON X-100 (C) at a concentration of zero or about 0.2 mM to about 10 mM. 14 H 22 O(C2H4O) n ) surfactant, chloride ions at a concentration of zero or about 200 mM to about 1,000 mM, a non-phenol-containing nonionic surfactant at a concentration of zero or about 0.2 mM to about 10 mM, glycerol at a concentration of zero or about 200 mM to about 800 mM, and NaSO at a concentration of zero or about 200 mM to about 800 mM.

[0115] Another embodiment of the disclosed composition may include the following components: BDEA at a concentration of about 75 mM to about 300 mM; Tris at a concentration of about 100 mM to about 300 mM; chloride ion at a concentration of about 200 mM to 1,000 mM; and TRITON X-100 at a concentration of about 0.2 to about 10 mM.

[0116] Another embodiment of the disclosed composition may include the following components: BDEA at a concentration of about 150 mM, Tris at a concentration of about 200 mM, KCl at a concentration of about 50 mM, TRITON X-100 at a concentration of about 1 mM, and NaCl at a concentration of about 800 mM.

[0117] Another embodiment of the disclosed composition may include the following components: BDEA at a concentration of about 75 mM to about 300 mM; Tris at a concentration of about 100 mM to about 300 mM; chloride ion at a concentration of about 200 mM to 1,000 mM; and a non-phenol-containing nonionic surfactant at a concentration of about 0.2 to about 5 mM, wherein the surfactant is Kolliphor P-407, Pluronic P-123, Pluronic L-121, Pluronic 31R1, Tetronic 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.

[0118] Another embodiment of the disclosed composition may include the following components: BDEA at a concentration of about 150 mM, Tris at a concentration of about 200 mM, KCl at a concentration of about 50 mM, a non-phenol-containing nonionic surfactant at a concentration of about 1 mM, and NaCl at a concentration of 800 mM, wherein the surfactant is KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.

[0119] In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component.

[0120] In embodiments, the present invention provides a compound of the formula: (HOCH2CH2)2N-CHR at a concentration of about 75 mM to about 300 mM: 1 -CHR 2 -CHR 3 -R4 (In the formula, R 1 , R 2 , R 3 , and R 4 are each independently -H, -CH3, -CH2CH3, or -CH(CH3)2, e.g., RR 1 , R 2 , R 3 , and R 4 and H), tris(hydroxymethyl)aminomethane (Tris) buffer at a concentration of zero or about 100 mM to about 300 mM, and TRITON X-100 (C) at a concentration of zero or about 0.2 mM to about 10 mM. 14 H 22 O(C2H4O) n ) surfactant, chloride ions at a concentration of zero or about 200 mM to about 1,000 mM, a non-phenol-containing nonionic surfactant at a concentration of zero or about 0.2 mM to about 10 mM, glycerol at a concentration of zero or about 200 mM to about 800 mM, and Na2SO4 at a concentration of zero or about 200 mM to about 800 mM.

[0121] In another embodiment of the disclosed composition, the following components are present: a compound of the formula: (HOCH2CH2)2N-CHR in a concentration of about 75 mM to about 300 mM; 1 -CHR 2 -CHR 3 -R 4 (In the formula, R 1 , R 2 , R 3 , and R 4 are each independently -H, -CH3, -CH2CH3, or -CH(CH3)2, e.g., RR 1 , R 2 , R 3 , and R 4 and each is H), Tris at a concentration of about 100 mM to about 300 mM, chloride ion at a concentration of about 200 mM to 1,000 mM, and TRITON X-100 at a concentration of about 0.2 to about 10 mM.

[0122] In another embodiment of the disclosed composition, the following components are present: 1 -CHR 2 -CHR 3 -R 4 (In the formula, R 1 , R 2 , R 3 , and R 4 are each independently -H, -CH3, -CH2CH3, or -CH(CH3)2, e.g., RR 1 , R 2 , R 3 , and R 4 and H), Tris at a concentration of about 200 mM, KCl at a concentration of about 50 mM, TRITON X-100 at a concentration of about 1 mM, and NaCl at a concentration of about 800 mM.

[0123] In another embodiment of the disclosed composition, the following components are present: a compound of the formula: (HOCH2CH2)2N-CHR in a concentration of about 75 mM to about 300 mM; 1 -CHR 2 -CHR 3 -R 4 ), where R 1 , R 2 , R 3 , and R 4 are each independently -H, -CH3, -CH2CH3, or -CH(CH3)2, e.g., RR 1 , R 2 , R 3 , and R 4 and each is H), Tris at a concentration of about 100 mM to about 300 mM, chloride ion at a concentration of about 200 mM to 1,000 mM, and a non-phenol-containing nonionic surfactant at a concentration of about 0.2 to about 5 mM, wherein the surfactant is KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ58, or TWEEN 20.

[0124] In another embodiment of the disclosed composition, the following components are present: 1 -CHR 2 -CHR 3 -R 4 (In the formula, R 1 , R 2 , R 3 , and R 4 are each independently -H, -CH3, -CH2CH3, or -CH(CH3)2, e.g., RR 1 , R 2 , R 3 , and R 4 wherein each is H), Tris at a concentration of about 200 mM, KCl at a concentration of about 50 mM, a non-phenol-containing nonionic surfactant at a concentration of about 1 mM, and NaCl at a concentration of about 800 mM, wherein the surfactant is KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.

[0125] In embodiments, the compositions of the present disclosure further comprise at least one of a surfactant, a liquid diluent, an ECL label, a binding reagent for a binding assay, a preservative, a biocide, an antifoaming agent, a perchlorate compound, a colorant, or a tracer chemical.

[0126] In embodiments, the present disclosure provides a composition consisting of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component.

[0127] In embodiments, the pH buffering component is tris(hydroxymethyl)-aminomethane (Tris).

[0128] In embodiments, the pH buffering component is tris(hydroxymethyl)-aminomethane (Tris), and the composition includes a surfactant. In embodiments, the surfactant includes a phenol ether. In embodiments, the surfactant is TRITON X-100. In embodiments, the surfactant does not include a phenol ether. In embodiments, the composition does not disrupt lipid bilayer membranes. In embodiments, the surfactant is KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the surfactant is TWEEN-20. In embodiments, the surfactant is PEG(18) tridecyl ether.

[0129] In embodiments, the composition of the present disclosure includes a surfactant that does not contain phenol ether. In embodiments, the composition does not disrupt lipid bilayer membranes. In embodiments, the surfactant is KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ58, or TWEEN 20. In embodiments, the surfactant is TWEEN-20. In embodiments, the surfactant is PEG(18) tridecyl ether. In embodiments, the pH buffering component is phosphate, HEPES, glycylglycine, borate, acetate, or citrate.

[0130] In embodiments, the ECL coreactant is BDEA. In embodiments, the ECL coreactant is DBAE. In embodiments, the ECL generated by the ECL label in the presence of the composition changes, on average, by less than 1% per degree Celsius over a temperature range of 18°C ​​to 30°C. In embodiments, the ECL is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode.

[0131] In embodiments of the compositions described herein, the pH is 6-9. In embodiments, the pH is 7-8. In embodiments, the pH is 7.6-7.9. In embodiments, the pH is about 7.8. In embodiments of the compositions described herein, the slope of the change in ECL with pH generated by the ECL label in the presence of the composition is less than 10% per pH unit. In embodiments, the ECL is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode.

[0132] In embodiments of the compositions described herein, the ionic component comprises NaCl. In embodiments, the ionic component comprises KCl. In embodiments, the ionic component comprises LiCl. In embodiments, the ionic component comprises chloride ions.

[0133] In embodiments, the compositions described herein include a liquid diluent, hi embodiments, the liquid diluent is water, and the compositions are substantially aqueous.

[0134] In embodiments of the compositions described herein, the concentration of the ECL coreactant is about 10 mM to about 800 mM. In embodiments, the concentration of the ECL coreactant is about 75 mM to about 300 mM. In embodiments, the concentration of the ECL coreactant is about 100 mM to about 150 mM. In embodiments of the compositions described herein, a change in the concentration of the ECL coreactant from 0.8 to 1.2 times its nominal value provides less than a 10% change in the ECL generated by the ECL label in the presence of the composition. In embodiments, the ECL is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode. In embodiments, the ECL from the ECL label is generated from an electrochemiluminescent ruthenium organometallic complex in proximity to a carbon-based electrode, and / or ECL in the absence of the ECL label is generated at a carbon-based electrode.

[0135] In an embodiment, the concentration of the pH buffering component is about 10 mM to about 800 mM. In an embodiment, the concentration of the pH buffering component is about 100 mM to about 300 mM. In an embodiment, the concentration of the pH buffering component is about 150 mM to about 250 mM.

[0136] In embodiments, the compositions described herein have an ionic strength greater than 0.3M. In embodiments, the compositions described herein have an ionic strength greater than 0.5M. In embodiments, the compositions described herein have an ionic strength greater than 0.8M. In embodiments, the compositions described herein have an ionic strength greater than 1.0M. In embodiments, the compositions comprise chloride ions, and the concentration of chloride ions is greater than about 0.25M. In embodiments, the compositions comprise chloride ions, and the concentration of chloride ions is greater than about 0.5M. In embodiments, the compositions comprise chloride ions, and the concentration of chloride ions is greater than about 0.75M. In embodiments, the compositions comprise chloride ions, and the concentration of chloride ions is greater than about 1.0M.

[0137] In embodiments of the compositions described herein, the pH buffering component is selected so that the composition provides at least a 20% reduction in ECL produced by the ECL label and / or at least a 20% reduction in background ECL produced in the absence of the ECL label, compared to the same composition containing phosphate as the pH buffering component.

[0138] In embodiments, non-specific binding (NSB) in immunoassays using compositions containing ionic components is lower compared to the same compositions without the ionic components.

[0139] In embodiments, the compositions described herein do not include a liquid diluent, and the compositions are provided in a dry form.

[0140] In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a surfactant. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a liquid diluent. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, a surfactant, and a liquid diluent.

[0141] In some embodiments, the compositions of the present disclosure further comprise at least one of an ECL label, a binding reagent for a binding assay, a preservative, a biocide, an antifoaming agent, a perchlorate compound, a colorant, a tracer chemical, a solid support, or a combination thereof.

[0142] In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a surfactant. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a liquid diluent. In embodiments, the present disclosure provides a composition consisting essentially of N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, a surfactant, and a liquid diluent.

[0143] In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, and an ionic component. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a surfactant. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, and a liquid diluent. In embodiments, the present disclosure provides a composition comprising N-butyldiethanolamine (BDEA) or 2-dibutylaminoethanol (DBAE), or both, a pH buffering component, an ionic component, a surfactant, and a liquid diluent.

[0144] In embodiments, the compositions described herein comprise about 75 mM to about 300 mM BDEA or DBAE, about 200 mM pH buffering component, and about 750 mM or more ionic component. In embodiments, the compositions comprise about 75 mM to about 300 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and about 1.0 mM surfactant. In embodiments, the compositions comprise about 75 mM to about 300 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 75 mM to about 300 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, about 1.0 mM surfactant, and an aqueous liquid diluent. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0145] In embodiments, the compositions described herein comprise about 150 mM BDEA or DBAE, about 100 mM to about 300 mM pH buffering component, and about 750 mM or more ionic component. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 100 mM to about 300 mM pH buffering component, about 750 mM or more ionic component, and about 1.0 mM surfactant. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 100 mM to about 300 mM pH buffering component, about 750 mM or more ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 100 mM to about 300 mM pH buffering component, about 750 mM or more ionic component, about 1.0 mM surfactant, and an aqueous liquid diluent. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0146] In embodiments, the compositions described herein comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, and about 500 mM to about 1500 mM ionic component. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 500 mM to about 1500 mM ionic component, and about 1.0 mM surfactant. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 500 mM to about 1500 mM ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 500 mM to about 1500 mM ionic component, about 1.0 mM surfactant, and an aqueous liquid diluent. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0147] In embodiments, the compositions described herein comprise about 75 mM to about 300 mM of an ECL coreactant selected from BDEA and DBAE, about 200 mM of a pH buffering component, and about 750 mM or more of an ionic component. In embodiments, the compositions comprise about 75 mM to about 300 mM of an ECL coreactant selected from BDEA and DBAE, about 200 mM of a pH buffering component, about 750 mM or more of an ionic component, and about 0.2 to about 10 mM of a surfactant. In embodiments, the compositions comprise about 150 mM of BDEA or DBAE, about 200 mM of a pH buffering component, about 750 mM or more of an ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 150 mM of BDEA or DBAE, about 200 mM of a pH buffering component, about 750 mM or more of an ionic component, about 0.2 to about 10 mM of a surfactant, and an aqueous liquid diluent. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0148] In embodiments, the compositions described herein comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, and about 750 mM or more ionic component. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and about 0.2 to about 10 mM TRITON X-100. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, about 0.2 to about 5.0 mM TRITON X-100, and an aqueous liquid diluent. In embodiments, the pH of the compositions is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0149] In embodiments, the compositions described herein comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, and about 750 mM or more ionic component. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and about 0.2 to about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, about 0.2 to about 5.0 mM non-phenol-containing non-ionic surfactant, and an aqueous liquid diluent. In embodiments, the pH of the compositions is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0150] In embodiments, the compositions described herein comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, and about 750 mM or more ionic component. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and about 0.2 to about 10 mM PEG(18) triethyl ether. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and an aqueous liquid diluent. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, about 0.2 to about 10 mM PEG(18) triethyl ether, and an aqueous liquid diluent. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0151] In embodiments, the compositions described herein comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, and about 750 mM or more ionic component. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and about 0.2 to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In embodiments, the compositions comprise about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, and an aqueous liquid diluent. In embodiments, the composition comprises about 150 mM BDEA or DBAE, about 200 mM pH buffering component, about 750 mM or more ionic component, about 0.2 to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether, and an aqueous liquid diluent. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0152] In embodiments, the compositions described herein comprise about 75 mM to about 300 mM BDEA, about 200 mM Tris, and about 750 mM or more chloride ions. In embodiments, the compositions comprise about 75 mM BDEA, about 0 or about 100 mM to about 300 mM Tris, and about 750 mM or more chloride ions. In embodiments, the compositions comprise about 75 mM to about 300 mM BDEA, about 200 mM Tris, and about 0 or about 500 mM to about 1500 mM chloride ions. In embodiments, the pH of the compositions is about 7.0 to about 8.0. In embodiments, the pH of the compositions is about 7.5. In embodiments, the pH of the compositions is about 7.8.

[0153] In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0154] In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing nonionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0155] In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM Kolliphor P-407, Pluronic P-123, Pluronic L-121, Pluronic 31R1, Tetronic 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM Kolliphor P-407, Pluronic P-123, Pluronic L-121, Pluronic 31R1, Tetronic 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5.0 mM Kolliphor P-407, Pluronic P-123, Pluronic L-121, Pluronic 31R1, Tetronic 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM Kolliphor P-407, Pluronic P-123, Pluronic L-121, Pluronic 31R1, Tetronic 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0156] In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 5.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0157] In one embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM to about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0158] In an embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0159] In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0160] In one embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0161] In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0162] In one embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0163] In one embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0164] In an embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0165] In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0166] In one embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0167] In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0168] In one embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether above its cmc. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0169] In one embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0170] In an embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0171] In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0172] In one embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0173] In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0174] In one embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0175] In one embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 150 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0176] In an embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0177] In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0178] In one embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0179] In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0180] In one embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0181] In one embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 200 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0182] In an embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0183] In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0184] In one embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0185] In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0186] In one embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0187] In one embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 300 mM BDEA, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0188] In embodiments, the compositions described herein comprise about 75 mM to about 300 mM DBAE, about 200 mM Tris, and about 750 mM or more chloride ions. In embodiments, the compositions comprise about 75 mM DBAE, about 0 or about 100 mM to about 300 mM Tris, and about 750 mM or more chloride ions. In embodiments, the compositions comprise about 75 mM to about 300 mM DBAE, about 200 mM Tris, and about 0 or about 500 mM to about 1500 mM chloride ions. In embodiments, the pH of the compositions is about 7.0 to about 8.0. In embodiments, the pH of the compositions is about 7.5. In embodiments, the pH of the compositions is about 7.8.

[0189] In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0190] In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and a non-phenol-containing nonionic surfactant at a concentration above its cmc.

[0191] In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM Kolliphor P-407, Pluronic P-123, Pluronic L-121, Pluronic 31R1, Tetronic 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition contains about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0192] In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 5.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0193] In one embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM to about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0194] In an embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0195] In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0196] In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0197] In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0198] In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0199] In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 75 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0200] In an embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0201] In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0202] In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0203] In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0204] In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of KOLLIPHOR P-407, PLURONIC P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0205] In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 100 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0206] In an embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0207] In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0208] In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0209] In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0210] In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0211] In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 150 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0212] In an embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0213] In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0214] In one embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0215] In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0216] In one embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0217] In one embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In another embodiment, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 200 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0218] In an embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, and about 500 mM to about 1500 mM chloride ion. In an embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, and about 750 mM or more chloride ion. In an embodiment, the pH of the composition is about 7.0 to about 8.0. In an embodiment, the pH of the composition is about 7.5. In an embodiment, the pH of the composition is about 7.8.

[0219] In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM TRITON X-100. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM TRITON X-100. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM TRITON X-100. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM TRITON X-100. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM TRITON X-100. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and TRITON X-100 at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0220] In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM of a non-phenol-containing non-ionic surfactant. In another embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM of a non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM non-phenol-containing non-ionic surfactant. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a non-phenol-containing non-ionic surfactant at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0221] In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the composition contains about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration above its cmc of KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0222] In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 5 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and about 10 mM Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether. In another embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ion, and a concentration of Kolliphor P-407, Pluronic P-123, or PEG(18) tridecyl ether above its cmc. In another embodiment, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8.

[0223] In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM to about 10 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 0.2 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 1.0 mM PEG(18) tridecyl ether. In one embodiment, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 5 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and about 10 mM PEG(18) tridecyl ether. In embodiments, the composition comprises about 300 mM DBAE, about 200 mM Tris, about 750 mM or more chloride ions, and PEG(18) tridecyl ether at a concentration above its cmc. In embodiments, the pH of the composition is about 7.0 to about 8.0. In embodiments, the pH of the composition is about 7.5. In embodiments, the pH of the composition is about 7.8. [Example]

[0224] The following examples are illustrative of some of the electrodes, plates, kits, and methods that fall within the scope of this disclosure. Of course, they are not to be construed as limiting the disclosure in any way. Those skilled in the art will be able to make numerous changes and modifications to this disclosure without undue experimentation.

[0225] Example I: Comparison of ECL coreactants ECL read buffers were prepared using three different ECL coreactants: tripropylamine (TPA), N-butyldiethanolamine (BDEA), and 2-dibutylaminoethanol (DBAE). Compared to TPA (vapor pressure = 2.9 mm Hg, boiling point = 156 °C), BDEA and DBAE have lower volatility (vapor pressure = 0.1 mm Hg and 1 mm Hg), higher boiling points (446 °C and 273-275 °C), lower flammability, higher water solubility, and less odor. The TPA-containing read buffer was MSD Read Buffer T (1X) (Meso Scale Discovery). BDEA and DBAE read buffers were prepared using the same formulation (125 mM coreactant, 200 mM Tris-HCl, 50 mM KCl, 0.1% (v / v) TRITON X-100, pH adjusted to 7.8 with HCl) except for replacing TPA with BDEA or DBAE. In addition, two additional read buffers were prepared that were similar to the TPA and BDEA read buffers described above except for the omission of the detergent (TRITON X-100). [ka]

[0226] ECL measurements were performed using an MSD MULTI-ARRAY streptavidin plate (Meso Scale Discovery). Each well of the plate incorporates a carbon ink-activated electrode supporting an immobilized layer of streptavidin. Before use, the wells were washed with phosphate-buffered saline containing TWEEN 20 (PBST). To measure the ECL signal from the ECL label, some wells of the plate were incubated with a solution containing a low concentration of bovine IgG labeled with both biotin NHS ester (to provide binding to the plate) and SULFO-TAG NHS ester (ECL label, Meso Scale Discovery). To measure the ECL background in the absence of label, some wells were incubated with the same solution except that the labeled bovine IgG was omitted. After incubation at room temperature with shaking to allow the labeled IgG to bind to the streptavidin-coated electrode, the wells were washed with PBST and one of the test read buffers was added. ECL was then measured using an ECL plate reader (MSD SECTOR Imager 6000, Meso Scale Discovery).

[0227] Figures 1A and 1B show that in detergent (TRITON X-100)-containing read buffers, BDEA, DBEA, and TPA all provided similar specific signals in the presence of ECL label (Figure 1A) and similar background signals in the absence of ECL (Figure 1B), indicating that BDEA and DBEA are viable alternatives to TPA. Consistent with the known benefits of detergents (especially aromatic ether-containing detergents such as TRITON X-100) for ECL in the presence of TPA-containing read buffers, the specific signal of the detergent-free TPA read buffer was substantially less than that of the detergent-containing TPA read buffer. In comparison, omission of detergent from the BDEA-containing read buffer had minimal effect on specific signal.

[0228] Figure 1C compares the surfactant-containing BDEA read buffer described in Figures 1A and 1B with a similar read buffer except that the Tris pH buffer component is replaced with phosphate as the buffer component. This figure shows that the use of phosphate as a buffer component results in a decrease in performance due to both a loss of specific signal and an increase in background signal.

[0229] Example II: Immunoassay using a BDEA-based read buffer The performance of a BDEA-based read buffer for multiplex sandwich immunoassays was compared with a conventional TPA-based read buffer using 10 multiplex assays from the MSD V-PLEX Proinflammatory Panel 1 kit (Meso Scale Discovery). The kit uses an MSD MULTI-ARRAY plate, where each well incorporates a screen-printed carbon ink electrode on the bottom and 10 arrays of capture antibodies against 10 analytes immobilized on the electrode. The assay was performed according to the kit instructions, except for the use of a BDEA-containing ECL read buffer. Briefly, (i) the plate was washed before adding samples; (ii) 50 μL of sample (containing calibration standards for each analyte diluted in kit assay diluent) was added to each well; (iii) the plate was incubated with shaking for 120 minutes to allow the analytes to bind to the capture antibodies and then washed to remove unbound sample; (iv) 25 μL of a mixture of 10 labeled detection antibodies for the 10 analytes (labeled with MSD SULFO-TAG NHS ester) in kit antibody diluent was added to each well; (v) the plate was incubated with shaking for 90 minutes to allow the detection antibodies to bind and capture the analytes and then washed to remove unbound detection antibodies; and (vi) 150 μL of ECL read buffer was added and ECL was measured on an MSD SECTOR Imager 6000 plate reader.

[0230] Figure 2A compares the signal measured using TPA and BDEA read buffers with optimized concentrations of each coreactant (the conventional TPA-containing read buffer specified by the kit (MSD Read Buffer T (2X)) and the BDEA-containing read buffer described in Example 1). The signal for the "Mid-Cal" sample, which contains a concentration of each analyte in the middle of the assay's quantitative range, and the signal for the "NSB" sample, which is the kit assay diluent with no added analyte, are shown. The figure shows the measured ECL signal for each analyte / array element and the ECL signal normalized to the signal measured using TPA read buffer. Each value represents the average of 240 replicates (5 assay plates x 48 replicates per plate) after removing outliers, in some cases, using a Grossman test with a 99% confidence interval. The figure also provides the average intra-plate coefficient of variation (CV) measured for each of the five plates (intra-plate CV), as well as the coefficient of variation of the average signal for each plate (inter-plate CV). The results show that at optimized concentrations, TPA and BDEA-based read buffers provide similar signals in the presence of analyte (normalized signals for Mid-Cal samples using BDEA-based read buffers range from 90% to 140%). The variability (expressed as CV) of the BDEA read buffer was, on average, similar to or slightly better than that of the TPA read buffer. Unexpectedly, when using BDEA buffer, the NSB sample signal (representing unwanted nonspecific background signal from nonspecific binding to the detection antibody capturing the array elements) was very high (the normalized NSB signal for one array element was close to 300%).

[0231] Figure 2B shows the results of repeating the experiment described above in Figure 2A, except that a further optimized BDEA read buffer was used and only four plates were tested per condition. The read buffer composition was as described in Example 1, but with a slightly higher BDEA concentration (150 mM) and also 800 mM NaCl. The addition of high salt levels had only a minor effect on the specific signal measured with the Mid-Cal samples (normalized signals ranged from approximately 75% to 125%), but had the unexpected ability to dramatically reduce nonspecific binding in the NSB samples (the highest normalized NSB signal was 152%, but most ranged from approximately 75% to 125%).

[0232] Example III: Robustness of the optimized BDEA read buffer This example shows the results of several experiments comparing the robustness of the optimized BDEA read buffer (shown above in Figure 2B) and the conventional TPA read buffer against different potential sources of environmental and compositional variation. Biotin and SULFO-TAG labeled bovine IgG (described in Example 1) bound to MSD streptavidin plates were used to generate a specific signal.

[0233] Figure 3A shows the effect of read buffer pH on the specific ECL signal (ECL) from bound labeled IgG and the background ECL measured in the absence of labeled IgG (background ECL). Results are shown as % change relative to the value measured at pH 7.8. The specific ECL signal measured over the pH 7.7-7.9 range was within 5% of the signal measured using the BDEA read buffer at pH 7.8 (filled circle symbols), but not in two conventional TPA-containing read buffers (MSD read buffer T (MSD T) 1X and 2X, diamonds and open circles, respectively). The slope of the change in specific ECL signal over the same pH range was approximately 7.4% per pH unit in the BDEA read buffer, compared to 161% per pH unit in read buffer T (1X) and 93% per pH unit in read buffer T (2X), demonstrating that the optimized BDEA formulation is much less sensitive to pH variations.

[0234] Figure 3B shows the effect of temperature on ECL generation in the presence of different read buffers. When the temperature was varied from approximately 18 °C to 30 °C and the optimized BDEA formulation was used as the read buffer, the specific signal (reported as % change from the signal at 21.8 °C) from biotin- and SULFO-TAG-labeled IgG on streptavidin plates remained within 10% of the value at 21.8 °C, whereas much larger changes were observed when using a conventional TPA read buffer (MSD read buffer T (2X)). The slope of the change in specific ECL signal over the temperature range tested was approximately 1.6% per °C for the BDEA read buffer compared to 3.9% per °C for read buffer T (2X), demonstrating that the optimized BDEA formulation is much less sensitive to temperature fluctuations.

[0235] Figure 3C shows the effect of coreactant concentration on ECL generation in the presence of different read buffers. The concentration of BDEA in the optimized BDEA read buffer formulation was varied from 0.8x (0.8X, 100 mM) to 1.4x (1.4X, 175 mM) of the nominal concentration (1X, 125 mM). Over these concentration ranges, there was little change in the measured specific ECL signal (remaining within 5% of the 1X condition). Similar signal and behavior were observed when TRITON X-100 was omitted from the formulation. The results indicate that the BDEA read buffer is relatively insensitive to changes in BDEA concentration resulting from differences in the BDEA raw material or due to manufacturing variability.

[0236] Figure 3D shows the effect of diluting or concentrating all components in the BDEA read buffer. The concentrations of all components in the optimized BDEA read buffer formulation were varied from 0.8x (0.8X) to 1.4x (1.4X) of the nominal concentration (1X). Over the range of 0.8X to 1.2X, the specific signal measured with the BDEA formulation remained within 10% of the 1X condition, whereas the variation in the specific signal measured with the TPA read buffer (MSD read buffer T 2X) was nearly twice that of the BDEA formulation. The results demonstrate that the BDEA read buffer is relatively insensitive to changes in dilution, such as user error in preparing the BDEA read buffer by reconstituting dry reagents or diluting liquid concentrates.

[0237] Figure 3E shows the effect of temperature on ECL generation in the presence of different read buffers (TPA read buffer, MSD read buffer T (2X), and BDEA formulation) and at various pHs ranging from 7.60 to 7.90. Figure 3F shows the % background change in ECL signal in the presence of TPA read buffer and BDEA formulation at different temperatures and pHs. The slope of the change in specific ECL signal over the temperature range tested at pH 7.80 is less than 1% per °C for BDEA read buffer compared to 6% per °C for read buffer T (2X), further demonstrating the robustness of the BDEA read buffer at different temperatures.

[0238] Figure 4 shows the effect of different voltage waveforms applied to streptavidin-coated electrodes of a streptavidin multiarray plate. The plot shows the specific signal (ECL) and background signal (background ECL) for four different voltage ramps (2.0 to 5.0 V, 2.5 to 4.5 V, 3.0 to 4.5 V, and 3.0 to 5.0 V) applied for different durations (1.5 to 12 seconds, depending on the ramp voltage). The ramp rate is determined by the ramp voltage and duration; for example, a ramp from 2.0 to 5.0 V over a 3-second duration would have a ramp rate of (5V - 2V) / 3 seconds = 1 V / second. The figure shows that for a given ramp, the background ECL increases with the duration of the read buffer for both the TPA and BDEA systems. In contrast, specific ECL increases with duration when using the TPA read buffer (MSD read buffer T 2X) but is relatively independent of duration when using the BDEA read buffer. The results indicate that the BDEA formulation is more tolerant to variations in ECL excitation conditions and that faster waveforms and measurements can be achieved with the BDEA read buffer without loss of sensitivity.

[0239] Example IV: BDEA Read Buffer with Alternative Surfactants Several BDEA-containing ECL read buffers were prepared with the composition of the optimized high-salt formulation of Example 2, except that the 1 mM TRITON X-100 component was replaced with 0.5 mM of various alternative milder surfactants (or the surfactant was omitted as a control). Figure 5A shows the surfactants tested in this experiment. Figure 5B shows that the addition of surfactant resulted in a significant reduction in the liquid-air surface tension of the read buffer (as determined by pendant droplet shape) compared to the read buffer without surfactant. For reproducible imaging into wells of a 96-well plate, a surface tension of less than 40 dynes / cm provided consistent meniscal shapes.

[0240] The efficiency of ECL generation in different read buffers was determined by measuring the specific signal from biotin- and SULFO-TAG-labeled bovine IgG bound to an MSD streptavidin plate, and the background signal was measured in the same system in the absence of labeled IgG (as described in Example 1). The specific ECL signal (Fig. 6A) and background ECL signal (Fig. 6B) were completely independent of detergent structure; all specific signals were within 10% and all background signals were within 10 ECL units of the values ​​measured in a read buffer containing TRITON X-100.

[0241] Different read buffer formulations were used to assay detergent-sensitive biological structures with lipid bilayer membranes (extracellular vesicles displaying the CD9 surface protein). MSD multi-array plates were prepared with a capture antibody against CD9 immobilized on integrated carbon ink actuation electrodes. Wells were incubated with samples containing extracellular vesicles. The wells were then washed to remove unbound sample and incubated with a solution containing a SULFO-TAG-labeled detection antibody against CD9 to bind the labeled detection antibody to the captured extracellular vesicles. The wells were then washed to remove unbound detection antibody, and read buffer was added. The plates were then analyzed immediately (time = 0) or incubated for 15 minutes before ECL measurement (time = 15) on an MSD SECTOR Imager 6000 ECL plate reader. Figure 7 shows the measured ECL signal. The figure shows that even at time = 0, exposing captured extracellular vesicles to a read buffer containing TRITON X-100 resulted in a near-complete loss of signal, likely due to vesicle lysis. In contrast, all other read buffers except BRIJ C10 gave signals within about 5% of each other at time = 0, although BRIJ C10 still gave signals within about 20%. Furthermore, all read buffers except those containing TRITON X-100, BRIJ C10, BRIJ S10, and BRIJ S20 exhibited less than 5% signal change when the plate was incubated in the read buffer for 15 minutes, demonstrating that the remaining detergents (KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-5-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, and TWEEN 20) do not disrupt captured extracellular vesicles or cause dissociation of the bound SULFO-TAG-labeled detection antibody.

[0242] The read buffer formulations were very stable, and accelerated stability testing of these formulations at 50 °C in the dark showed minimal or no significant changes in performance in most cases, except for the TWEEN 20, PLURONIC 31R1, and TETRONIC 701 formulations, which produced approximately 25% less specific signal after 220 days of exposure to this temperature. The formulations were also robust to changes in surfactant concentration. Figure 8 shows that varying the concentration of PEG(18) tridecyl ether from 0.5 to 4 mM had no significant effect on the specific ECL signal in the CD9 extracellular vesicle assay or on the stability of the captured binding reaction product after 15 minutes of incubation in the read buffer.

[0243] Example V: Immunoassay Off-Rates In antibody screening experiments with different capture and detection antibodies for various analytes, read buffers containing TPA (MSD T2X) and BDEA were tested. Figures 9A-9D show the average % ECL loss, which is an indication of the off-rate between the antibody and the analyte (e.g., between the capture antibody and / or detection antibody and the analyte), for antibody screens of sRange (Figure 9A), IL-9 (Figure 9B), Kim-1 (Figure 9C), and MIG (Figure 9D). In each of the antibody screens, the BDEA read buffer reduced the % ECL loss compared to the TPA read buffer, demonstrating that the BDEA read buffer can reduce the antibody-analyte off-rate compared to the TPA read buffer.

[0244] Figures 10A-10C further demonstrate the improvement in antibody-analyte off-rates with BDEA compared to TPA read buffer in multiplex panel experiments using the analytes IL-13 (Figure 10A), MDC (Figure 10B), and TNF-β (Figure 10C). In each of the experiments, the BDEA read buffer showed a reduced antibody-analyte off-rate, while the nonspecific background signal (NSB) with the BDEA read buffer was either comparable or improved compared to the TPA read buffer.

[0245] Example VI: Extracellular vesicle assay TPA read buffer (MSD T1X) and BDEA read buffer containing varying concentrations of TRITON X-100 were tested for performance in an intact extracellular vesicle (EV) assay. The change in EV assay signal with each buffer type and TRITON X-100 concentration was measured.

[0246] The results are shown in Figure 11A. For MSD T1X, assay performance improved as the TRITON X-100 concentration was decreased from 0.1% to 0.01%. As the TRITON X-100 concentration was further decreased from 0.01% to 0%, the performance of the MSD T1X assay decreased. BDEA read buffer with 0.1% TRITON X-100 had poor assay performance, while BDEA read buffer with 0% TRITON X-100 had the best assay performance of all buffer types and TRITON X-100 concentrations tested.

[0247] A BDEA read buffer containing a detergent that does not dissolve EVs was tested for assay performance variability. Titration curves were generated using known concentrations of CD81+ EVs for two different lots of non-TRITON BDEA read buffer. The results are shown in Figure 11B. The two tested lots of non-TRITON BDEA read buffer had very similar titration curves, indicating low lot-to-lot variability in performance.

[0248] 7. Incorporating References The present disclosure is not limited in scope by the specific embodiments described herein. Indeed, various modifications of the present disclosure in addition to those described herein will become apparent to those skilled in the art from the foregoing description and accompanying figures. Such modifications are intended to be included within the scope of the following claims. Various publications are cited herein, the disclosures of which are incorporated by reference in their entireties.

[0249] The described embodiments and examples of the present disclosure are intended to be illustrative rather than limiting, and are not intended to represent all embodiments or examples of the present disclosure. Moreover, while basic novel features of the present disclosure as applied to various specific embodiments thereof have been shown, described, and pointed out, it will be understood that various omissions, substitutions, and changes in the form and details of the illustrated devices, as well as their operation, may be made by those skilled in the art without departing from the spirit of the present disclosure. For example, all combinations of these elements and / or method steps that perform substantially the same function in substantially the same way to achieve the same results are expressly intended to be within the scope of the present disclosure. Furthermore, it should be recognized that structures and / or elements and / or method steps shown and / or described in connection with any disclosed form or disclosed embodiment may be incorporated into any other disclosed or described or proposed form or embodiment as a matter of general design choice. Moreover, various changes and modifications can be made without departing from the spirit or scope of the disclosure, as set forth in the following claims, both literally and in equivalents recognized in law.

Claims

1. (i) 2-dibutylaminoethanol (DBAE), (ii) a pH buffering component, (iii) an ionic component, and (iv) a surfactant; the concentration of DBAE is about 100 mM to about 150 mM; the concentration of the ionic component is from about 200 mM to about 1400 mM; composition.

2. (i) 2-dibutylaminoethanol (DBAE), (ii) a pH buffering component, (iii) an ionic component, and (iv) a surfactant; the concentration of DBAE is about 100 mM to about 150 mM; the concentration of the ionic component is from about 200 mM to about 1400 mM; composition.

3. 3. The composition of claim 1, wherein the pH buffering component is not a phosphate.

4. The composition of any one of claims 1 to 3, wherein the pH buffering component is Tris.

5. The composition according to any one of claims 1 to 4, wherein the surfactant is TRITON X-100.

6. The composition of any one of claims 1 to 4, wherein the surfactant is KOLLIPHOR P-407, PLURONIC P-123, PLURONIC L-121, PLURONIC 31R1, TETRONIC 701, 2,4,7,9-tetramethyl-d-decyne-4,7-diol ethoxylate, PEG(18) tridecyl ether, BRIJ L4, BRIJ 58, or TWEEN 20.

7. The composition of any of claims 1 to 6, wherein the ionic component comprises chloride ions.

8. The composition of any of claims 1 to 7, wherein the composition has an ionic strength of greater than about 0.3M.

9. In one or more containers, (a) 2-dibutylaminoethanol (DBAE), (b) a pH buffering component; and (c) an ionic component; and (d) a surfactant, the concentration of DBAE is about 100 mM to about 150 mM; the concentration of the ionic component is from about 200 mM to about 1400 mM; kit.

10. 1. A method for producing a composition, comprising: (i) 2-dibutylaminoethanol (DBAE), (ii) a pH buffering component; and (iii) an ionic component; and (iv) a surfactant, the concentration of DBAE is about 100 mM to about 150 mM; the concentration of the ionic component is from about 200 mM to about 1400 mM; method.

11. A composition according to any one of claims 1 to 8 or a kit according to claim 9; (a) an assay device; (b) assay consumables; (c) additional assay reagents; (d) an assay sample; or a combination thereof.

12. 1. A method for producing ECL, comprising: (a) an electrode; (i) a composition according to any one of claims 1 to 8, and (ii) contacting with an ECL label; (b) applying a voltage to the electrodes; (c) producing ECL.

13. 1. A method for measuring the amount of ECL label, comprising: (a) an electrode; (i) a composition according to any one of claims 1 to 8, and (ii) contacting with said ECL label; (b) applying a voltage to the electrodes; (c) generating ECL; (d) measuring the ECL; (e) determining the amount of the label from the measured ECL.

14. 1. A method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, comprising: (a) contacting a binding reagent immobilized on an electrode with a labeled binding reagent comprising an ECL label; (b) forming a binding complex on the electrode comprising the immobilized binding reagent and the labeled binding reagent; (c) contacting the binding complex on the electrode with a composition according to any one of claims 1 to 8; (d) applying a voltage to the electrodes in the presence of the composition; (e) generating ECL; (f) measuring the ECL to determine the amount of the binding complex on the electrode.

15. 1. A method for measuring the amount of a binding complex comprising a binding reagent linked to an ECL label, comprising: (a) contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label; (b) forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent; (c) contacting the binding complex on the particles with a composition according to any one of claims 1 to 8; (d) collecting the particles on an electrode; (e) applying a voltage to the electrodes in the presence of the composition; (f) generating ECL; (g) measuring the ECL to determine the amount of the binding complex on the electrode.

16. 1. A method for determining the amount of an analyte, comprising: (a) contacting a binding reagent immobilized on an electrode with a labeled binding reagent comprising an ECL label and the analyte (or a sample comprising the analyte); (b) forming a binding complex on the electrode comprising the immobilized binding reagent and the labeled binding reagent; (c) contacting the binding complex on the electrode with a composition according to any one of claims 1 to 8; (d) applying a voltage to the electrodes in the presence of the composition; (e) generating ECL; (f) measuring said ECL to determine said amount of said analyte.

17. 1. A method for determining the amount of an analyte, comprising: (a) contacting a binding reagent immobilized on a particle with a labeled binding reagent comprising an ECL label and an analyte (or a sample comprising said analyte); (b) forming a binding complex on the particle comprising the immobilized binding reagent and the labeled binding reagent; (c) contacting the binding complex on the particles with a composition according to any one of claims 1 to 8; (d) collecting the particles on an electrode; (e) applying a voltage to the electrodes in the presence of the composition; (f) generating ECL; (g) measuring the ECL to determine the amount of the analyte.

18. 1. A composition comprising: (i) 2-dibutylaminoethanol (DBAE); (ii) a pH buffering component; (iii) an ionic component; and (iv) a surfactant; the concentration of DBAE is about 100 mM to about 150 mM; the concentration of the ionic component is from about 200 mM to about 1400 mM; A composition wherein the pH buffering component is not a phosphate.

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