Lateral flow device for detecting target analytes and related methods thereof

Abstract

The present disclosure relates to a lateral flow device for detecting target analytes in a biological sample. The lateral flow device may include a strip including: (i) a sample-receiving region disposed adjacent the proximal end; (ii) at least one porous membrane; (iii) a flow control region having a water-soluble polymer and a sugar alcohol; (iv) a filter region having a non-target antigen; and (v) a first detection region having a second analyte-specific binding agent. The lateral flow device described herein improves sensitivity and accuracy of a lateral flow assay.

Description

Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT LATERAL FLOW DEVICE FOR DETECTING TARGET ANALYTES AND RELATED METHODS THEREOFRELATED PATENT APPLICATIONS

[0001] This patent application claims the benefit of U.S. Provisional Patent Application No.63 / 777,631 filed on March 25, 2025, entitled Lateral Flow Device For Detecting Target Analytes And Related Methods Thereof, naming Phillip Hartzog and Jun Lu as inventors, and designated by Attorney Docket No. 056718-1474240. The entire content of the foregoing application is incorporated herein by reference, including all text, tables and drawings.FIELD

[0002] The present disclosure generally relates to devices and methods for detecting target analytes. More specifically, the present disclosure generally relates to an improved lateral flow device that provides improved sensitivity and accuracy of detection of target analytes in a sample.BACKGROUND

[0003] A conventional lateral flow device (LFD) is often a type of diagnostic test used to detect the presence, absence, or amount of a target substance (e.g., a virus, bacteria, or other biomarker) in a sample. Conventional LFDs often work by allowing a liquid sample to flow through a membrane strip that contains reagents that react and / or bind with a target substance, providing a visible result, usually in the form of a line or a color change. These devices are commonly used for rapid testing, for example, for pregnancy (e.g., detecting human chorionic gonadotropin) or pathogen testing (e.g., COVID-19 antigen tests). Lateral flow tests are often valued for their simplicity, ease of use, and the ability to deliver quick result. A traditional LFD test is typically performed at home, in a clinic, or in another non-lab oratory setting because traditional LFDs often do not require any specialized equipment to interpret the results.

[0004] Presented herein is an improved lateral flow device and methods of use thereof.SUMMARY

[0005] In some embodiments, presented herein is a lateral flow device for detecting a target analyte in a sample, the device including: a strip having a proximal end and a distal end, the strip comprising one or more of (i) a sample-receiving region disposed adjacent the proximalAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT end; (ii) at least one porous membrane; (iii) a flow control region comprising a water-soluble polymer and / or a sugar alcohol; (iv) a filter region comprising a non-target antigen; and (v) a first detection region comprising a second analyte-specific binding agent. In some embodiments, the lateral flow device further comprises colloidal gold nanoparticles comprising a polydopamine coating and a first analyte-specific binding agent that binds specifically to the target analyte.

[0006] In some embodiments, presented herein is a method for detecting a target drug in a sample. The method comprises providing a lateral flow device as described herein; contacting a sample with the sample-receiving region of the lateral flow device; and determining a presence, absence, or amount of gold nanoparticles in the first detection region.

[0007] Additional embodiments will become apparent upon consideration of the detailed description.BRIEF DESCRIPTION OF THE DRAWINGS

[0008] The drawings illustrate embodiments of the technology and are not limiting. Reference to figures and / or features of a figure in this document are not limiting. For clarity and ease of illustration, the drawings are not made to scale and, in some instances, various aspects may be shown exaggerated or enlarged to facilitate an understanding of particular embodiments.

[0009] FIG. 1 shows a top view of an embodiment of a lateral flow device 100 comprising strip 101 and optional substrate 106. Strip 101 comprises, from the proximal end 200 to distal end 250, one or more of a sample-receiving region 118, particle region 119, buffering region 120, flow control region 125, filter region 130, first detection region 140, intervening region 135, second detection region (e.g., control region) 145, and absorption pad 150. In this embodiment, strip 101 comprises a porous membrane 110 located between the samplereceiving region 118 and absorption pad 150. The porous membrane 110 comprises a proximal end 220 and a distal end 240.

[0010] FIG. 2 shows a side view of the lateral flow device shown in Fig. 1.DETAILED DESCRIPTION

[0011] Described herein is an improved lateral flow device for detecting target analytes in a sample.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0012] A sample can be obtained from, or derived from a suitable subject. A sample can be isolated or obtained directly from a subject or part thereof. In some embodiments, a sample is obtained indirectly from an individual or medical professional. A sample can be any specimen that is isolated or obtained from a subject or part thereof. In some embodiments, a sample comprises a bodily fluid obtained from a subject. In some embodiments, a sample comprises an extract or lysate of a tissue or cells. Non-limiting examples of a sample include blood or a blood product (e.g., serum, plasma, platelets, buffy coats, or the like), umbilical cord blood, amniotic fluid, interstitial fluid, cerebrospinal fluid, spinal fluid, lavage fluid (e.g., lung, gastric, peritoneal, ductal, ear, arthroscopic), urine, sputum, saliva, nasal mucous, prostate fluid, lavage, semen, lymphatic fluid, bile, tears, sweat, breast milk, breast fluid, a liquid biopsy, as well as lysates or extracts obtained from cells (blood cells, lymphocytes, placental cells, stem cells, bone marrow derived cells, embryo or fetal cells), feces, tissues, the like or combinations thereof. In some embodiments, a sample is blood or a blood product, e.g., serum. A sample may be processed prior to contact with a lateral flow device described herein. For example, a sample may be partially purified, filtered, concentrated, and / or diluted prior to contact with a lateral flow device described herein.

[0013] The term “subject” refers to an animal, typically a mammalian animal. In some embodiments a subject is a mammal. Non-limiting examples of mammals include humans, non-human primates (e.g., apes, gibbons, chimpanzees, orangutans, monkeys, macaques, and the like), domestic animals (e.g., dogs and cats), farm animals (e.g., horses, cows, goats, sheep, pigs) and experimental animals (e.g., mouse, rat, rabbit, guinea pig). In some embodiments, a subject is a primate. In some embodiments, a subject is a human.

[0014] In some embodiments, a lateral flow device (e.g., 100) described herein is configured to detect a presence or absence of, and / or determine an amount of, a target analyte in a sample.

[0015] In some embodiments, a target analyte is any molecule of interest found in a sample, non-limiting examples of which include proteins, nucleic acids, carbohydrates, fatty acids, organic small molecules, inorganic molecules, steroids, pathogens, drugs, metabolites, and portions thereof. A target analyte may be naturally occurring or synthetic. In some embodiments, a target analyte is a drug, or metabolite thereof, non-limiting examples of which include prescription drugs and over-the-counter drugs, including barbiturates, depressants, antidepressants, analgesics, hallucinogens, inhalants, opioids, stimulants, benzodiazepines, cannabinoids, NSAIDs, ACE inhibitors, biologies, etc. and the like. In some embodiments, a target analyte is a drug of abuse or a metabolite thereof. A drug of abuse may include naturallyAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT occurring drugs, synthetic drugs, non-naturally occurring drugs, or metabolites thereof. Nonlimiting examples of a drug of abuse include depressants and opioids including heroin, codeine, fentanyl, hydrocodone (dihydrocodeinone), hydromorphone, meperidine, methadone, morphine, oxycodone or oxymorphone; stimulants including amphetamines, phenethylamines, amphetamine sulfate, methamphetamine, dextroamphetamine, levoamphetamine, lisdexamfetamine, atomoxetine, methylphenidate, dexmethylphenidate, oxymetazoline, pseudoephedrine, phenylephrine, benzodiazepines including alprazolam, chlordiazepoxide, diazepam, lorazepam or triazolam, barbiturates including phenobarbital, pentobarbital, methohexital, secobarbital, butobarbital or butalbital, as well as eszopiclone, zaleplon and zolpidem, the like and metabolites thereof. In some embodiments, a target analyte is fentanyl or norfentanyl.

[0016] In some embodiments, a lateral flow device described herein has a limit of detection for a target analyte in a sample of at least 40 ng / ml, at least 5 ng / ml, at least 1 ng / ml, at least 0.5, at least 0.1, or at least 0.01 ng / ml. In some embodiments, a lateral flow device has a limit of detection for a target analyte in a sample of less than 20 ng / ml, less than 10 ng / ml, less than 5 ng / ml, less than 1 ng / ml, less than 0.1 ng / ml or less than about 0.01 ng / ml. In some embodiments, a lateral flow device has a limit of detection for a target analyte in a range of 50 ng / ml to about 0.01 ng / ml, or 20 ng / ml to about 0.01 ng / ml. In some embodiments, a lateral flow device has a limit of detection for a target analyte (e.g., a drug, small molecule, or drug of abuse) of about 5 ng / ml, about 1 ng / ml, about 0.1 ng / ml or about 0.01 ng / ml. In some embodiments, a lateral flow device has a limit of detection for fentanyl or norfentanyl of about 5 ng / ml, about 1 ng / ml, about 0.1 ng / ml or about 0.010 ng / ml.

[0017] In some embodiments, a lateral flow device has a limit of detection for a target analyte in a sample of less than 100 nM, less than 50 nM, less than 20 nM, less than 10 nM, less than 1 nM, less than 0.1 nM or less than 0.01 nM. In some embodiments, a lateral flow device has a limit of detection for a target analyte in a range of 50 nM to about 0.01 nM, about 15 nM to about 1 nM, about 10 nM to about 0.1 nM, or about 10 nM to about 0.01 nM. In some embodiments, a lateral flow device has a limit of detection for a target analyte (e.g., a drug, small molecule, or drug of abuse) of about 15 nM, about 10 nM, about 1 nM, about 0.1 nM or about 0.01 nM. In some embodiments, a lateral flow device has a limit of detection for fentanyl or norfentanyl of about 10 nM, about 1 nM, about 0.1 nM or about 0.01 nM.

[0018] Accordingly, the lateral flow device described herein often increases detection on the order of 2 to 1000-fold over conventional lateral flow devices.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0019] FIG. 1 shows an exemplary embodiment of a lateral flow device 100. A lateral flow device (e.g., 100) is often configured to detect the presence, absence, and / or amount of a target analyte in a sample. A lateral flow device 100 may comprise a strip 101, and optionally a substrate 106. In some embodiments the strip 101 is attached to a substrate 106. In some embodiments, a substrate 106 comprises a non-porous, non-absorbent and / or liquid impervious surface. In some embodiments, a lateral flow device or a strip does not comprise a substrate. For example, a strip 101 can be independent of a substrate of a lateral flow device 100. A sample can be applied onto, or delivered to, a sample-receiving region 118 of strip 101. For example, in embodiments where a lateral flow device is worn by a subject, or contacted with the skin of a subject, a sample can be delivered to the sample-receiving region using a suitable method, e.g., by capillary action.

[0020] A lateral flow device or strip of a lateral flow device can be any suitable shape or size. In some embodiments, a strip is a linear strip. In some embodiments, a strip is a rectangular strip, circular strip, an oval strip, or another shape that allows movement of a sample from one side of a strip to another side of a strip. In some embodiments, a strip is substantially flat. In some embodiments, a strip is a rectangular shape and has a length in a range of 50 mm to 200 mm, 50 mm to 100 mm, or about 60 mm to about 80 mm. In some embodiments, a strip is a rectangular shape and has a width in a range of about 3 mm to 30 mm, 4 mm to 20 mm, or about 4 mm to about 6 mm.

[0021] In some embodiments, a strip 101 comprises at least one porous membrane 110. In some embodiments, a strip comprises one porous membrane or a single contiguous porous membrane. In some embodiments, a strip comprises 2, 3, 4, 5, 6, 7, 8, 9 or 10 porous membranes. In some embodiment, a strip comprises two or more porous membranes arranged consecutively from a distal end of the strip to a proximal end of the strip such that each porous membrane of the strip is adjacent to and / or in contact with another porous membrane of the strip. In certain embodiments, one porous membrane of a strip may overlap a portion of another porous membrane of the strip. In some embodiments, one or more porous membranes of a strip comprise the same material or different materials. The one or more porous membranes of a strip are often configured to allow a sample to migrate (e.g., by capillary action and / or diffusion) from a proximal end of a strip to a distal end of the strip. In some embodiments, one or more porous membranes of a strip are often configured to allow a sample to migrate (e.g., by capillary action and / or diffusion) from a sample-receiving end of a strip toward a distal end of the strip.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0022] In some embodiments, a porous membrane 110 of a lateral flow device 100 comprises a suitable material that allows and / or promotes migration of a sample from a proximal portion of a strip to a distal portion of a strip, non-limiting examples of which include polysaccharides (e.g., cellulose material, paper (e.g., whatman paper), and cellulose derivatives such as cellulose acetate and nitrocellulose), polyether sulfone, polyethylene, nylon, polyvinylidene fluoride, polyester, polypropylene, silica, inorganic materials uniformly dispersed in a porous polymer matrix with a polymer such as vinyl chloride, vinyl chi ori de-propylene copolymer, and vinyl chloride-vinyl acetate copolymer, such as inactivated alumina, diatomaceous earth, MgSC>4, cotton) and synthetic (e.g., nylon or rayon) fabrics, porous gels, such as silica gel, agarose, dextran, and gelatin, polymeric films such as polyacrylamide, and the like. In some embodiments, a porous membrane 110 comprises nitrocellulose.

[0023] In some embodiments, a porous material of a lateral flow device comprises a suitable pore size. Non-limiting examples of a suitable pore size includes pores having a an absolute or average diameter in a range of 0.01 to about 20 microns (pm), 0.1 to about 20 pm, 0.1 to about 20 pm, 0.1 to about 15 pm, 0.1 to about 10 pm, 0.1 to about 8 pm, 0.1 to about 5 pm, 0.25 to about 5 pm, and intermediate ranges thereof.

[0024] In certain embodiments, strip 101 comprises a proximal end 200 and a distal end 250. In some embodiments, a strip comprises one or more portions or regions (e.g., 110, 118, 119, 120, 125, 130, 135, 140, 145 & 150) each comprising a proximal end and a distal end. A proximal end of a portion or region of a strip is the end of the portion or region that is closest to the proximal end 200 of strip 101. A distal end of a portion or region of a strip is the end of the portion or region that is closest to the distal end 250 of strip 101.

[0025] The terms “proximal to” and “distal to” as used herein refer to a location of a region or portion of an LFD or strip relative to another region or portion of the LFD or strip. For example, if a first region is proximal to a second region on a strip, the first region is located between the proximal end 200 of strip 101 and a proximal end of the second region. For example, regions 118, 119, 120 and 125 in FIG. 1 are all located proximal to region 130. In yet another example, if a first region is distal to a second region on a strip, the first region is located between the distal end 250 of strip 101 and a distal end of the second region. For example, regions 130, 135, 140, 145 and 150 in FIG. 1 are all located distal to region 125.

[0026] A lateral flow device or strip of a lateral flow device often comprises one or more regions, non-limiting examples of which include a sample-receiving region, sample pad, buffer region, flow control region, filter region, detection region, control region and absorbent region.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT In some embodiments, a region of a strip comprises a width equal to the width of a strip and a length (e.g., measured from a proximal end to a distal end of the region) in a range of about 1-20 mm, about 1-10 mm, about 1-3 mm, or about 1 mm. In some embodiments, a region of a strip may be separated from, adjacent to, or overlapping (partially or completely) with one or more other regions of the strip.

[0027] In some embodiments, strip 101 comprises a sample-receiving region 118. In some embodiments, a sample-receiving region 118 of strip 101 is located adjacent to a proximal end 200 of the strip. In some embodiments, a sample-receiving region 118 of strip 101 is disposed adjacent to a proximal end 200 of the strip. In some embodiments, a sample-receiving region of a strip is proximal to one or more of regions 119, 120, 125, 130, 135, 1400, 145, and 150 of a strip. A sample receiving-region is configured to receive a sample (e.g., a liquid or aqueous sample). For example, a sample-receiving region is configured to receive a volume of sample in a range of 0.1 to 5000 pl. In some embodiments, a sample-receiving region is configured to receive a volume of sample in a range of 0.1 to 2500 pl, 0.1 to 1000 pl, 0.1 to 500 pl, 1 to 500 pl, 10 to 500 pl, or intermediate ranges therein.

[0028] In some embodiments, a sample-receiving region 118 comprises a suitable porous membrane or pad. In some embodiments, the sample-receiving region 118 includes a sample pad comprising a porous material, non-limiting examples of which include nitrocellulose, cellulose, polyester, porous polyethylene, sponge, paper, glass fiber filter paper, silica, cotton fiber, glass fiber, woven glass fibers, chopped glass, glass, the like, derivatives thereof, and combinations thereof. In some embodiments, a sample pad is treated with, and / or comprises a suitable buffer and / or blocking agent. In some embodiments, a sample pad comprises tri s(hydroxymethyl)aminom ethane and / or borate. In some embodiments, a sample-receiving region 118 comprises a porous membrane or pad and / or a non-porous surface.

[0029] In some embodiments, a sample-receiving region 118 comprises a porous membrane or pad that is attached to, or disposed on, a non-porous surface. Non-limiting examples of a non-porous surface include plastic (e.g., polystyrene, polyethylene, polypropylene, polyvinyl chloride, and the like), glass, borosilicate, ceramic, the like, derivatives thereof, and combinations thereof. In some embodiments, a sample-receiving region comprises a well. In some embodiments, a well of a sample-receiving region is located above the plane of a strip. In some embodiments, a well of a sample-receiving region includes a mouth and sidewalls to guide a sample to the sample-receiving region. In some embodiments, a lateral flow deviceAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT comprises a substrate (e.g. , housing) comprising a well located at or above the sample-receiving region.

[0030] In some embodiments, a porous material or pad of a sample-receiving region comprises a suitable pore size, for example configured to filter residual components (e.g., cells) from a sample. Non-limiting examples of a suitable pore size includes pores having a an absolute or average diameter in a range of about 0.01 to about 30 microns (pm), about 0.1 pm to about 20 pm, about 15 pm to about 20 pm, about 15 pm to about 20 pm, about 1 pm to about 20 pm, about 3 pm to about 20 pm, about 3 pm to about 10 pm, about 3 pm to about 8 pm, about 10 pm to about 12 pm, and intermediate ranges thereof. In some embodiments, a porous material of a lateral flow device or strip has an average pore size of about 3 pm to about 20 pm.

[0031] In some embodiments, a strip, or one or more regions or portion thereof comprises one or more buffering regions (e.g. , 120) or suitable buffers. In certain embodiments, a samplereceiving region 118 and / or sample pad comprises a buffering region 120. In some embodiments, a buffering region is located on the distal end of a sample-receiving region 118. In some embodiments, a strip 101 comprises a buffering region 120 located adjacent to a sample-receiving region or particle region (e.g., 119). In some embodiments, a buffering region 120 is located proximal to a flow control region 125, a filter region 130, a first detection region 140, and a second detection region 145. In some embodiments, a buffering region 120 is located proximal to a blocking region 125 and / or proximal to a filter region 130.

[0032] A buffering region often comprises one or more suitable buffers. Buffers are often disposed in or on a buffering region and are often dried on a lateral flow device, or portion thereof. A buffer is often configured to maintain a pH of a sample in a range of about 5 to about 8, about 6.5 to about 8, or about 7 to about 8. For example, a buffer often maintains a pH of a sample at a pH of about 5, 6, 7, 7.5, or 8. In some embodiments, a sample and a buffer are mixed prior to contacting a sample with a sample-receiving region or sample pad of a lateral flow device described herein.

[0033] In some embodiments, a buffering region comprises a suitable surfactant. In certain embodiments, a suitable surfactant is a non-ionic detergent, non-limiting examples of which include Tween-20, Tween-80, NP-40, and Triton X-100.

[0034] In some embodiments, a buffering region comprises detectable particles.

[0035] In some embodiments, a buffering region is about 2 mm to about 20 mm long, or about 2 to about 10 mm long. In certain embodiments, a buffering region spans a total widthAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT of a strip. In some embodiments, a buffering region has a surface area of about 3 mm2to about 40 mm2.

[0036] In some embodiments, a lateral flow device or strip does not comprise detectable particles. In such an embodiment, detectable particles are often mixed with a sample prior to contacting the particles and / or sample with a lateral flow device.

[0037] In some embodiments, a region or portion of a lateral flow device or strip comprises detectable particles. In some embodiments, a particle region (e.g., 119) comprises detectable particles. In certain embodiments, a particle region is separate and distinct from other regions on a strip. In some embodiments, a sample-receiving region or pad comprises detectable particles. In some embodiments, a buffer region, blocking region, or flow control region comprises detectable particles. In some embodiments, a region of a lateral flow device or strip that includes detectable particles (e.g., 119) is located proximal to a detection region (e.g., a first or second detection region). In some embodiments, a region of a strip that includes detectable particles is located proximal to any of regions 120, 125, 130, 135, 140, 145, and / or 150, if present, and as illustrated in the example of FIG. 1. In some embodiments, a region of a strip that includes detectable particles is located proximal to regions 140, 135, 145, and / or 150, if present, and distal to regions 118, 120, 125 and / or 130. In certain embodiments, a particle region is located distal to a sample-receiving region and proximal to a buffer region. In some embodiments, a sample-receiving region (e.g., 118) or sample pad comprises detectable particles. In some embodiments, a distal portion of a sample-receiving region (e.g., 118) or sample pad comprises detectable particles. In certain embodiments, a buffering region (e.g., 120) comprises a plurality of detectable particles. In some embodiments, detectable particles of a lateral flow device are located between a sample-receiving region and a blocking region, and / or between a sample-receiving region and a filter region of a strip.

[0038] In some embodiments, a region of a strip that comprises detectable particles, or a particle region (e.g., 119) is about 1 mm to about 20 mm or about 1 mm to about 10 mm long. In certain embodiments, a region of a strip that comprises detectable particles, or a particle region (e.g., 119) spans a total width of a strip. In some embodiments, a region of a strip that comprises detectable particles, or a particle region (e.g., 119) has a surface area of about 3 mm2to about 40 mm2.

[0039] Detectable particles are often attached to one or more binding agents that are configured to bind specifically to a target analyte or antigen. In some embodiments, detectable particles are nanoparticles. In some embodiments, detectable particles have an average size inAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT a range of about 1 to about 500 nanometers (nm), 10 nm to 500 nm, 20 nm to 250 nm, 1 nm to 150 nm, about 20 nm to about 100 nm, about 10 to 50 nm, about 10 nm to 49 nm, or about 10 nm to 45 nm. In some embodiments, detectable particles are coated.

[0040] In some embodiments, detectable particles comprise magnetic particles, polymeric particles, carbon-based nanoparticles, metal oxide nanoparticles, metal nanoparticles, the like or combinations thereof.

[0041] In some embodiments, one or more detectable particles comprise a suitable detectable label, non-limiting examples of which include a chromogen, an enzyme, a catalyst, a fluorescent compound, chemiluminescent compound, a dye, a metal, the like, and combinations thereof.

[0042] In some embodiments, a detectable particle comprises a suitable metal, non-limiting examples of which include silver, copper, platinum, gold, zinc, cadmium, cobalt, titanium, lead, iron, the like or combinations thereof. In some embodiments, detectable particles are gold particles or colloidal gold particles. In some embodiments, detectable particles are colloidal gold nanoparticles.

[0043] In some embodiments, colloidal gold particles have an absolute or average particle size (i.e., diameter) less than 200 nm, less than 100 nm, less than 50 microns, less than 48 nm, less than 45 nm, or less than 40 nm. In some embodiments, colloidal gold particles of a lateral flow device as described herein have an average particle size in a range of 0.1 to 500 nm, 1 nm to 500 nm, 1 nm to 250 nm, 1 nm to 150 nm, 20 nm to 100 nm, 1 nm to 45 nm, or 10 nm to 45 nm. An average or absolute diameter of colloidal gold particles is often determined prior to coating or conjugation.

[0044] In some embodiments, one or more detectable particles (e.g., colloidal gold particles), or a portion thereof, comprise a suitable coating. As described herein, a coating of a detectable particle is often configured to reduce non-specific binding, and / or increase the mass or size of some or all of the detectable particles used in a lateral flow device. Non-limiting examples of a coatings include polysaccharides, peptides, polydopamine, polymeric amines, chitosan, alginate, polyphenol -based coating (e.g., tannic acid and catechol), the like or combinations thereof. In some embodiments, one or more detectable particles comprise a polydopamine coating. Detectable particles (e.g., colloidal gold particles) can be coated using a suitable method. In some embodiments, colloidal gold particles are coated by oxidative selfpolymerization of dopamine on the surface of the colloidal gold particles. One or more, or a portion of detectable particles used in, or with, a lateral flow device may be completely coated,Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT partially coated, or not coated. In some embodiments, a ratio of polydopamine coated gold nanoparticles to uncoated gold nanoparticles is in a range from about 10: 1 to about 1:10, about 5:1 to about 1:5, about 2:1 to about 1:2, about 1:2 to about 1:4, or about 2:1 to about 1:5. In some embodiments, a ratio of polydopamine coated gold nanoparticles to uncoated gold nanoparticles is at least about 1 :2. In certain embodiments, an amount of polydopamine coated gold nanoparticles used in, or with, a lateral flow device, is about 100% to 5% of a plurality of gold nanoparticles, or about 75% to 10%, 50% to 15%, or about 30% to about 20% of a plurality of gold nanoparticles.

[0045] In some embodiments, detectable particles (e.g., colloidal gold nanoparticles; e.g., coated or uncoated gold nanoparticles) comprise a binding agent configured to specifically bind to a target analyte (i.e., an analyte-specific binding agent). A binding agent can be attached to one or more detectable particles (e.g., gold particles) using a suitable method.

[0046] The term "specifically binds" refers to a binding agent that binds to a target analyte in preference to binding other molecules or other peptides as determined by, for example, a suitable in vitro assay (e.g., an Elisa, Immunoblot, Flow cytometry, and the like). A specific binding interaction discriminates over non-specific binding interactions by about 2-fold or more, often about 10-fold or more, and sometimes about 100-fold or more, 1000-fold or more, 10,000- fold or more, 100,000-fold or more, or 1,000,000-fold or more.

[0047] In some embodiments, a binding agent comprises or consists of a suitable antibody, an antibody fragment and / or an antigen binding portion thereof (e.g., a binding fragment). An antibody can refer to a natural antibody, polyclonal antibody, monoclonal antibody, recombinant antibody, a chimeric antibody, an antibody binding fragment (e.g., an antigen binding portion of an antibody), a CDR-grafted antibody, a humanized antibody, a human antibody, or portions thereof. In some embodiments a binding agent comprises or consists of one or more suitable antigen binding portions of an antibody, non-limiting examples of which include Fab, Fab', F(ab')2, Fv fragment, single-chain Fv (scFv), diabody (Dab), synbody, the like and / or a combination or portion thereof. In some embodiments a binding agent comprises TandAbs, aptamers, nanobodies, BiTEs, SMIPs, DARPins, DNLs, affibodies, Duocalins, adnectins, fynomers, Kunitz Domains AlbudAbs, DARTs, DVD-IG, Covx-bodies, peptibodies, scFv-Igs, SVD-Igs, dAb-Igs, Knob-in-Holes, triomAbs, the like or combinations thereof. In some embodiments, a binding agent comprises a single-chain polypeptide comprising one or more antigen binding portions of an antibody.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0048] In some embodiments, one or more detectable particles are colloidal gold particles coated (completely or partially) with polydopamine, wherein the particles comprise a binding agent that binds specifically to a target analyte (e.g., fentanyl).

[0049] In some embodiments, a lateral flow device or strip comprises a flow control region (e.g., 125). In certain embodiments, a flow control region comprises one or more reagents that modulate the flow, diffusion, and / or migration of a sample, reagents, and / or detectable particles on a strip. In some embodiments, a flow control region is configured to slow the flow of a sample and reagents (e.g., detectable particles) through a porous membrane of a strip. Applicants have determined that a flow control region can increase specific binding of binding agents to a target analyte and improve accuracy, sensitivity, and / or limit of detection of a lateral flow assay.

[0050] In some embodiments, a flow control region 125 of a strip 101 is located adjacent to a sample-receiving region 118. In some embodiments, a flow control region 125 is distal to a sample-receiving region 118, particle region 119, and / or a buffering region 120. In some embodiments, the flow control region 125 can be directly adjacent to the sample-receiving region 118, particle region 119, and / or buffering region 120. In some embodiments, the flow control region 125 can be directly adjacent to buffering region 120. In some embodiments, a flow control region 125 of a strip is proximal to regions 130, 140, 135, 145, and 150. In some embodiments, a portion of a flow control region 125 can be disposed on a sample-receiving region 118, particle region 119, buffering region 120, and / or filter region 130.

[0051] In some embodiments, a flow control region 125 is located on a region of at least one porous membrane 110 of a strip. In some embodiments, a flow control region 125 of a strip is located adjacent to a proximal end 220 of the at least one porous membrane 110. In some embodiments, a flow control region (e.g., 125), a filter region (e.g., 130), a detection region (e.g., 135), or combinations thereof, are disposed on a porous membrane (e.g., 110).

[0052] In some embodiments, a flow control region is about 1-20 or about 1-10 mm long. In certain embodiments, a flow control region spans a total width of a strip. In some embodiments, a flow control region has a surface area of about 3 mm2to about 40 mm2.

[0053] In some embodiments, a flow control region comprises a water-soluble polymer, nonlimiting examples of which include polyvinyl alcohol (PVA), polylactic acid, polyethylene glycol, alginate, and chitosan. In some embodiments, a water-soluble polymer comprises polyvinyl alcohol. In some embodiments, a water-soluble polymer has a degree of hydrolysis in a range of about 50% (mol%) to about 99%, about 70% to about 99%, about 80% to aboutAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT 92%, or about 85% to about 90%. In some embodiments, a flow control region comprises PVA having a degree of hydrolysis of about 87% to about 89%.

[0054] In some embodiments, a water-soluble polymer has a molecular weight (g / mmol) in a range of about 5,000 to about 70,000 g / mol, about 10,000 to about 50,000 g / mol, about 10,000 to about 30,000 g / mol, or about 10,000 to about 25,000 g / mol. In some embodiments, a water-soluble polymer has a molecular weight (g / mmol) in a range of about 13,000 to about 23,000 g / mol.

[0055] In some embodiments, a flow control region comprises about 1-100 ug, about 1-50 ug, about 1 to 25 ug, about 1 to 15 ug, about 1 to 10 ug, or about 5-10 ug of a water-soluble polymer. In certain embodiments, a flow control region comprises about 5, 6, 7, 8 or 9 ug of a water-soluble polymer. In certain embodiments, a flow control region comprises about 5, 6, 7, 8 or 9 ug of PVA. In certain embodiments, a flow control region comprises about 0.1 to about 5 ug / mm2, or about 0.1 to about 2 ug / mm2of a water-soluble polymer.

[0056] In some embodiments, a strip or region thereof (e.g., a flow control region) comprises a suitable sugar alcohol. In certain embodiments, a suitable sugar alcohol comprises a solubility in a range of about 10 g / 100 ml to about 30 g / 100 ml, or about 15 g / 100 ml to about 25 g / 100 ml in water. In certain embodiments, a sugar alcohol comprises a solubility of less than about 30 g / 100 ml, or less than about 25 g / 100 ml of water. Non-limiting examples of a sugar alcohol include sorbitol, lactitol, mannitol, maltitol, erythritol, arabitol, ribitol, galactitol, and rhamnitol. In some embodiments, a flow control region comprises mannitol.

[0057] In some embodiments, a region of a strip or flow control region comprises about 1-100 ug, about 1-50 ug, about 1 to 25 ug, about 1 to 20 ug, or about 10 to 20 ug of a sugar alcohol. In certain embodiments, a flow control region comprises about 15, 16, 17, 18, 19 or 20 ug of a sugar alcohol. In certain embodiments, a flow control region comprises about 15, 16, 17, 18, 19 or 20 ug of mannitol. In certain embodiments, a flow control region comprises about 1-5 ug / mm2, or about 3-4 ug / mm2of a sugar alcohol.

[0058] In some embodiments, a lateral flow device or flow control region of a lateral flow device does not comprise a sugar alcohol.

[0059] In some embodiments, a flow control region comprises a water-soluble polymer and a sugar alcohol. In some embodiments, a flow control region comprises polyvinyl alcohol and mannitol. In some embodiments, a ratio of a sugar alcohol to a water-soluble polymer is in a range from about 1:1 to 4:1, from about 1:2 to 4:1, or from about 3:1 to 4:1. In some embodiments, the flow control region comprises at least 20% (wt. % / unit area), at least 25%,Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT at least 30%, at least 35%, at least 40%, at least 50%, at least 55%, or at least 60% of a sugar alcohol. In some embodiments, the flow control region comprises from about 20% to 65% sugar alcohol, for example, about 25% to 55%, about 25% to 50%, about 20% to 40%, or about 25% to 45%, based on the total dried weight of solute in the flow control region. In some embodiments, the flow control region comprises at least 30% water-soluble polymer, for example, at least 35%, at least 40%, at least 50%, at least 55%, at least 60%, at least 65%, or at least 70%, based on the total dried weight of solute in the flow control region. In some embodiments, the flow control region comprises from about 30% to 75% water-soluble polymer, for example, about 35% to 70%, about 40% to 70%, about 45% to 65%, or about 40% to 60%, based on the total dried weight of solute in the flow control region.

[0060] In some embodiments, a flow control region is formed by depositing an aqueous solution on a flow control region of a porous membrane, the solution comprising a water-soluble polymer in an amount of about 0.5-10% or about 1.5-3%, a sugar alcohol in an amount of about 1 to 20%, or about 3-6%. In some embodiments, the solution comprises PVA in an amount of about 0.5-10% or about 1.5-3%, and mannitol in an amount of about 1 to 20%, or about 3-6%. In some embodiments, the solution comprises a blocking agent (e.g., BSA) in an amount of about 0.5-15%, or about 2-6%. Reagents of a flow control region are often dried on a porous membrane of a strip.

[0061] In some embodiments, a strip, one or more regions of a strip or a flow control region comprises one or more of a blocking agent, a buffer and / or a salt. A blocking agent is often a relatively inert substance, or a mixture of substances, that can block non-specific binding of target analytes, proteins, detectable particles, and binding agents to themselves, to each other, or to a portion of a lateral flow device. Non-limiting examples of blocking agents include peptides, proteins, amino acids, macromolecular blocking agents, gelatin, carbohydrates, surfactants (e.g., Tween or Tween 20), various polymers (e.g., polyvinylpyrrolidone), milk (e.g., non-fat milk), fatty acids, the like, extracts or digests thereof, and combinations thereof. In some embodiments, a blocking agent comprises a protein. In some embodiments, a blocking agent comprises albumin (e.g., bovine serum albumin). In some embodiments, blocking agent is dried and / or disposed on a flow control region, or other region of a strip.

[0062] In certain embodiments, a strip comprises a dried blocking agent in an amount of about 1-100, about 1-50 or about 5-20 ug per strip, or about 0.5-5 ug / mm length of a strip.

[0063] In some embodiments, a lateral flow device or strip comprises a filter region (e.g., 130). In certain embodiments, a filter region is configured to capture unbound binding agentsAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT (e.g., binding agents conjugated to detectable particles not yet bound to a target analyte), and immobilize those binding agents so that they do not migrate to a detection region. In some embodiments, a filter region comprises one or more immobilized antigens (i.e., non-target antigens) that are different than a target analyte. In some embodiments, a filter region comprises a non-target antigen. A non-target antigen of a filter region often comprises an epitope similar to an epitope present on a target analyte. In some embodiments, a non-target antigen of a filter region comprises an epitope that a target-analyte specific binding agent is known to cross-reacts with. In some embodiments, a non-target antigen of a filter region comprises a similar structure to a target analyte. In some embodiments, a binding agent attached to a detectable particle can cross-react and / or bind to a non-target antigen of a filter region.

[0064] In certain embodiments, a non-target antigen of a filter region is not a binding agent, is not an antibody, and / or does not comprise a portion of an antibody. In certain embodiments, a non-target antigen of a filter region does not comprise protein A or protein G.

[0065] In some embodiments, a non-target antigen is any naturally occurring or synthetic molecule, non-limiting examples of which include a protein, nucleic acid, carbohydrate, sugar, lipid, fatty acid, small molecule, organic small molecule, inorganic molecule, steroid, pathogen, drug, the like, metabolites thereof, and portions thereof. In some embodiments, a non-target antigen is a drug, or metabolite thereof, non-limiting examples of which include prescription drugs and over-the-counter drugs, including barbiturates, amphetamines, depressants, antidepressants, analgesics, hallucinogens, inhalants, opioids, stimulants, benzodiazepines, cannabinoids, NSAIDs, ACE inhibitors, biologies, etc. and the like. In some embodiments, a non-target antigen is a drug of abuse or a metabolite thereof. In some embodiments, wherein a target analyte is fentanyl, a non-target antigen is an amphetamine, an analog thereof, a derivative thereof or metabolite thereof. In some embodiments, a non-target antigen is not fentanyl. In some embodiments, a non-target antigen is phenethylamine.

[0066] In some embodiments, where a target analyte and corresponding analyte-specific binding agent have been identified for use in a lateral flow device described herein, a non-target antigen can be selected by screening the analyte-specific binding agent for cross-reactive binding to antigens that share a similar structure with the target analyte. For example, where a target analyte is a protein, the analyte-specific binding agent can be screened for binding to other proteins in the same family or class, or to other proteins that share acid sequences having high homology or identity with the target analyte, or a portion thereof. Various alignmentAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT algorithms and / or programs may be used to determine percent identity or percent homology, including FASTA, BLAST, or ENTREZ. FASTA and BLAST, which are available as a part of the GCG sequence analysis package (University of Wisconsin, Madison, Wis.). ENTREZ is available through the National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Md. Where a target analyte is a small molecule or drug, non-target antigens can readily be identified that are in the same family or class of small molecules and / or that share highly similar structures with the target analyte. Screening for cross-reactivity can be performed using a standard immunoassay, e.g., an ELISA assay.

[0067] In some embodiments, a non-target antigen is irreversibly attached to, or immobilized on, a strip of a lateral flow device described herein. In some embodiments, a non-target antigen is irreversibly attached to, or immobilized on, at least one porous membrane of a strip. In some embodiments, a non-target antigen is irreversibly attached to, or immobilized on, a filter region of a strip.

[0068] A non-target antigen can be irreversibly attached to, or immobilized on, a porous membrane or strip using a suitable method. In some embodiments, a non-target antigen, (e.g., a protein) is directly immobilized on a membrane of a strip using a suitable method. In some embodiments, a non-target antigen is attached to an anchor moiety which is then irreversibly attached to, or immobilized on, a membrane of a strip or lateral flow device using a suitable method. In certain embodiments, an anchor moiety is any molecule that can be attached to a non-target antigen and immobilized on a non-porous membrane of a strip. In some embodiments, an anchor moiety comprises a protein (e.g., BSA). For example, where a target analyte is fentanyl or norfentanyl, and a non-target antigen is an amphetamine, the amphetamine can be conjugated to BSA, which is then immobilized on a nitrocellulose membrane prior to blocking.

[0069] In some embodiments, a filter region 130 can be disposed on a region of the at least one porous membrane 110. The filter region 130 can be located adjacent to a proximal end 220 of the at least one porous membrane 110. In some embodiments, the filter region 130 is adjacent to the flow control region 125 on the at least one porous membrane 110. In some embodiments, a filter region 130 of a strip is proximal to one or more of regions 140, 135, 145, and 150 of a strip. The filter region 130 can be disposed between a flow control region 125 and a first detection region 140 of the strip 101. In some embodiments, the sample-receivingAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT region 118, particle region 119, buffering region 120, and / or flow control region 125 are located proximal to filter region 130.

[0070] In some embodiments, a lateral flow device or strip comprises a detection region (e.g., 140). In some embodiments, a porous membrane of a strip comprises a detection region. In some embodiments, a lateral flow device or strip comprises two or more detection regions (e.g., 140 & 145). In some embodiments, a lateral flow device or strip comprises a first detection region and a second detection region. In some embodiments, a first detection region is configured to bind to a target analyte and a second detection region (often located distal to the first detection region) is a control region. In some embodiments, a first detection region is configured to bind to a first target analyte, a second detection region is configured to bind to a second target analyte, and a third detection region (often located distal to the first and second detection regions) is a control region. In some embodiments, an intervening region 135 comprises one or more detection regions.

[0071] In some embodiments, a detection region comprises one or more binding agents configured to bind specifically to an antigen (e.g., a target analyte). One or more binding agents of a detection regions are often irreversibly attached or immobilized on a porous membrane of a strip or lateral flow device described herein. In certain embodiments, a detection region comprises one or more binding agents (e.g., a plurality of binding agents). Binding agents of a particular detection region are often configured to specifically bind to the same antigen.

[0072] In some embodiments, a first detection region comprises a binding agent that specifically binds to a target analyte. A binding agent of a first detection region is often different than a binding agent that is attached to a detectable particle. For example, a binding agent immobilized in a detection region often binds to an epitope on a target analyte that is different (e.g., non-competitive) than an epitope of the target analyte that is recognized by the binding agent attached to a detectable particle.

[0073] In some embodiments, a second detection region is a control region. In certain embodiments, a control region comprises an immobilized binding agent configured to bind an antigen known to be present in a sample, where the antigen is not the target analyte. In some embodiment, a control region comprises an immobilized binding agent configured to bind the binding agent attached to a detectable particle. For example, where a detectable particle comprises a first binding agent configured to bind to a target analyte, a control region may comprise an immobilized second binding agent configured to bind to the first binding agent. In some embodiments, an immobilized binding agent of a control region does not bind to aAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT target analyte. In some embodiments, a binding agent of a control region binds specifically to a non-binding portion (e.g., an Fc region, or heavy chain constant region) of another binding agent. For example, where a first binding agent that is attached to a detectable particle is a mouse antibody, a second immobilized binding agent of a control region can be a goat antimouse antibody.

[0074] As shown in the embodiment of FIG. 1, strip 101 or porous membrane 110 comprises a first detection region 140. A first detection region is often distal to a sample-receiving region 118, particle region 119, buffering region 120, flow control region 125, and / or filter region 130, when present. In some embodiments, a first detection region 140 is proximal to a control region (e.g., second detection region 145). A control region is often distal to a first detection region. In some embodiments, a second detection region that is configured to detect a second target analyte is located between a first detection region and control region.

[0075] In some embodiments, a strip includes an absorbent pad or absorption pad (e.g., 150). In some embodiments, an absorption pad 150 is located adjacent to a distal end 250 of strip 101. In some embodiments, an absorption pad 150 is distal to a sample-receiving region 118, particle region 119, buffering region 120, flow control region 125, filter region 130, and detection regions 140 and 145. The absorption pad 150 generally receives a sample moving through strip 101, and may help promote capillary action and diffusion of a sample through a porous membrane 110. The absorption pad 150 absorbs and removes excess sample or buffer to prevent disturbing the results.

[0076] In some embodiments, an absorption pad comprises a suitable porous material.

[0077] In some embodiments, a lateral flow device may comprise a substrate (e.g., 106). In some embodiments a strip is attached to a substrate. In some embodiments, a lateral flow device or a strip does not comprise a substrate. For example, a strip can be independent of a substrate of a lateral flow device.

[0078] In some embodiments, the strip 101 may be disposed on or attached to a substrate 106. A substrate may be liquid impervious, non-porous, and / or non-absorbent. A substrate can comprise a suitable liquid impervious, non-porous, and / or non-absorbent material, nonlimiting example of which include glass; and polymeric materials such as polystyrene, polypropylene, polyester, polybutadiene, polyvinylchloride, polyamide, polycarbonate, epoxide, methacrylate, and polymelamine. In some embodiments, a lateral flow device comprises a housing that partially encloses a strip of a lateral flow device. A housing may comprise a sample application region configured to allow sample application to a sample-Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT receiving region, and in some embodiments, a viewing region that allows visual inspection of one or more detection region.

[0079] In some embodiments, most reagents found on one or more regions of a strip (e.g., sample-receiving region, sample pad, buffer region, flow control region and filter region, when present) are reversibly attached to a strip of a lateral flow device described herein. For example, in certain embodiments, detectable particles, buffers, blocking agents, sugar alcohols, water-soluble polymers, alcohols, and other reagents found in, for example, regions 118, 119, 120, 125, and 130, are reversibly attached to a strip, and such reagents are capable of diffusion and / or migration within a strip upon contacting a liquid. Reagents can be reversibly attached to a strip using a suitable method. In some embodiments, reagents are irreversibly attached to a strip by drying said reagents on the strip. Often, reagents on a strip are completely or partially dried and / or desiccated prior to contact with a sample.

[0080] In some embodiments, certain reagents, for example binding agents (e.g., binding agents attached to a detection region), are irreversibly attached to and / or immobilized on a strip using a suitable method. A reagent that is irreversibly bound to, or immobilized on a strip is often fixed in position, and is not expected to migrate, diffuse or move upon contact with a liquid, buffer or sample.

[0081] Provided herein, in some embodiments, is a method of performing a lateral flow assay to detect a presence, absence, or amount of a target analyte in a sample. In some embodiments, a method comprises (a) providing a lateral flow device described herein and (b) contacting a sample with a sample-receiving region or pad of the lateral flow device, and (c) determining a presence, absence, or amount of the detectable particles in a first detection region of the lateral flow device. In some embodiments, a determination of the presence of detectable particles in the first detection region indicates a presence or amount of the target analyte in the sample and / or a determination of the absence of detectable particles in the first detection region indicates the absence of the target analyte in the sample.

[0082] In some embodiments, the lateral flow device comprises detectable particles, where the detectable particles comprise a binding agent configured to specifically bind to a target analyte of interest. In some embodiments, the method comprises contacting the sample with detectable particles prior to (a), where the detectable particles comprise a binding agent configured to specifically bind to a target analyte of interest. In some embodiments, detectable particles comprise colloidal gold nanoparticles. In some embodiments, the target analyte is fentanyl or norfentanyl.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0083] In some embodiments, a kit comprises a lateral flow device described herein. In some embodiments, a kit comprises colloidal gold particles comprising a plurality of binding agents configured to specifically bind to a target analyte of interest. In some embodiments, a kit comprises a pipette. In some embodiments, a kit comprises a collection device. In some embodiments, a kit comprises a blood draw device. In some embodiments, the kit includes a fluid transfer device. In some embodiments, a fluid transfer device can be a vial, container, pipette, collection container, or the like. In certain embodiments, a sample collector such as a finger prick needle or a pipette for depositing a sample may be included.Further Embodiments

[0084] Al. A lateral flow device for detecting a target analyte in a sample, the device comprising: a strip having a proximal end and a distal end, the strip comprising: (i) a samplereceiving region disposed adjacent the proximal end; (ii) at least one porous membrane; (iii) a flow control region comprising a water-soluble polymer and / or a sugar alcohol; (iv) a filter region comprising a non-target antigen; and (v) a first detection region comprising a second analyte-specific binding agent.

[0085] Al.l: The device of embodiment Al, further comprising a plurality of detectable particles comprising a first analyte-specific binding agent.

[0086] Al. 2: The device of embodiment Al.l, wherein the detectable particles comprise a coating.

[0087] A2: The device of embodiment Al.l or Al.2, wherein the detectable particles comprise coated and / or uncoated colloidal gold nanoparticles.

[0088] A2.1: The device of any one of embodiments Al.2 or A2, wherein some or all of the detectable particles comprise a polydopamine coating.

[0089] A3 : The device of embodiment Al, Al .1 or A2, wherein the flow control region, the filter region, and the first detection region are disposed on the at least one porous membrane.

[0090] A4: The device of any one of embodiments A1-A3, wherein the at least one porous membrane comprises nitrocellulose.

[0091] A5: The device of any one of embodiments A2-A4, wherein the colloidal gold nanoparticles are disposed on a distal portion of the sample-receiving region.

[0092] A6: The device of any one of embodiments A2-A4, wherein the colloidal gold nanoparticles are disposed on the at least one porous membrane.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0093] A7: The device of any one of embodiments A1-A6, wherein the sample-receiving region comprises a sample pad.

[0094] A8: The device of embodiment A7, wherein the sample-receiving region or the sample pad comprises a non-porous, or non-absorbent material.

[0095] A9: The device of embodiment A7, wherein the sample-receiving region or the sample pad comprises a porous material.

[0096] A10: The device of embodiment A7, wherein the sample-receiving region or the sample pad comprises an absorbent material.

[0097] Al 1 : The device of any one of embodiments A7-A10, wherein the sample-receiving region or the sample pad comprises silica, cotton fiber, or woven glass fibers.

[0098] A12: The device of any one of embodiments A7-A11, wherein the colloidal gold nanoparticles are disposed on a portion of the sample-receiving region or the sample pad

[0099] A12.1: The device of any one of embodiments A2-A12, wherein the colloidal gold particles a disposed on a distal portion of the sample-receiving region or sample pad, and adjacent to the flow control region.

[0100] A13: The device of any one of embodiments A1-A12.1, wherein the samplereceiving region comprises a buffer.

[0101] A14: The device of embodiment A13, wherein the buffer is configured to maintain a pH of a sample fluid in a range of 5-8.

[0102] A15: The device of any one of embodiments A1-A14, wherein the sample-receiving region comprises a surfactant.

[0103] A16: The device of any one of embodiments A2-A15, wherein the colloidal gold nanoparticles comprise uncoated or non-dopamine coated gold nanoparticles.

[0104] Al 7: The device of embodiment Al 6, wherein a ratio of the poly dopamine coated gold nanoparticles to the non-dopamine coated gold nanoparticles is in a range of 5: 1 to 1 :5.

[0105] Al 8: The device of any one of embodiments A2-A17, wherein the first analytespecific binding agent specifically binds to a first portion or first epitope of the target analyte.

[0106] A19: The device of any one of embodiments A2-A18, wherein the first analytespecific binding agent comprises an antibody or antigen binding portion thereof.

[0107] A20: The device of any one of embodiments A1-A19, wherein the water-soluble polymer comprises a polymer having a degree of hydrolysis of at least 50%.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0108] A21: The device of any one of embodiments A1-A20, wherein the water-soluble polymer is selected from the group consisting of polyvinyl alcohol, polylactic acid, polyethylene glycol, alginate, and chitosan.

[0109] A21: The device of any one of embodiments Al -21, wherein the water-soluble polymer is polyvinyl alcohol.

[0110] A23: The device of any one of embodiments A1-A22, wherein the sugar alcohol has a solubility of less than 25 g / 100 ml of water.[OHl] A24: The device of any one of embodiments A1-A23, wherein the sugar alcohol is one or more of sorbitol, lactitol, mannitol, maltitol, erythritol, arabitol, ribitol, galactitol, and rhamnitol.

[0112] A25: The device of any one of embodiments A1-A24, wherein the sugar alcohol is mannitol.

[0113] A26: The device of any one of embodiments A1-A25, wherein the water-soluble polymer is polyvinyl alcohol and the sugar alcohol is mannitol.

[0114] A27: The device of any one of embodiments A1-A26, wherein the flow control region comprises a blocking agent.

[0115] A28: The device of embodiment A27, wherein the blocking agent comprises a protein.

[0116] A29: The device of embodiment A28, wherein the blocking agent comprises bovine serum albumin.

[0117] A30: The device of any one of embodiments A1-A29, wherein the flow control region comprises 2.0 to 4.0 wt. % water-soluble polymer, 3.0 to 10.0 wt. % sugar alcohol, and 2.0 to 8.0 wt. % blocking agent.

[0118] A31: The device of any one of embodiments A1-A30, wherein a ratio of the sugar alcohol to the water-soluble polymer is in a range from 1 : 1 to 4: 1.

[0119] A32: The device of any one of embodiments A1-A31, wherein the flow control region further comprises methanol.

[0120] A33: The device of any one of embodiments A1-A32, the flow control region is separate and distinct from the sample-receiving region and the first detection region.

[0121] A34: The device of any one of embodiments A1-A33, wherein the flow control region is proximal to the first detection region and distal to the sample-receiving region.

[0122] A34.1: The device of any one of embodiments A1-A34, wherein the non-target antigen is irreversibly attached to, or immobilized on, the at least one porous membrane.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0123] A35: The device of any one of embodiments A1-A34.1, wherein the non-target antigen is attached or conjugated to an anchor moiety.

[0124] A36: The device of embodiment A35, wherein the anchor moiety comprises a bead or particle.

[0125] A > . The device of embodiment A35 or A36, wherein the anchor moiety comprises a protein.

[0126] A38: The device of embodiment A37, wherein the anchor moiety comprises bovine serum albumin.

[0127] A39: The device of any one of embodiments A1-A38, wherein the non-target antigen is reversibly attached to the at least one porous membrane.

[0128] A40: The device of any one of embodiments A1-A39, wherein the filter region is proximal to the first detection region and distal to the sample-receiving region.

[0129] A41: The device of any one of embodiments A1-A40, wherein the filter region comprises at least 50 % less bovine serum albumin than the flow control region.

[0130] A42: The device of any one of embodiments A1-A41, wherein the filter region comprises less than 0.10 wt. % conjugated bovine serum albumin.

[0131] A43: The device of any one of embodiments Al -42, wherein the target analyte is a drug, a drug of abuse, a synthetic drug, a non-naturally occurring drug or a metabolite thereof.

[0132] A44: The device of embodiment A43, wherein the drug of abuse is a phenethylamine.

[0133] A45: The device of embodiment A43 or A44, wherein the drug of abuse comprises fentanyl, an analog, a derivative or metabolite thereof.

[0134] A46: The device of embodiment A45, wherein the metabolite of fentanyl is norfentanyl.

[0135] A47: The device of any one of embodiments A1-A46, wherein the target analyte is different than the non-target antigen.

[0136] A48: The device of any one of embodiments A43-A47, wherein the non-target antigen is a drug of abuse, a synthetic drug, a non-naturally occurring drug or a metabolite thereof.

[0137] A49: The device of any one of embodiments A43-A48, wherein the non-target antigen comprises a phenethylamine.

[0138] A50: The device of any one of embodiments A43-A49, wherein the non-target antigen comprises an amphetamine, an analog, a derivative or metabolite thereof.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0139] A51 : The device of any one of embodiments A1-A50, wherein the at least one porous membrane comprises one porous membrane.

[0140] A52: The device of any one of embodiments A1-A51, wherein a portion of the at least one porous membrane comprises a cellulose derivative.

[0141] A53: The device of embodiment A52, wherein the cellulose derivative comprises a pore size in a range of 5 to 20 micrometers.

[0142] A54: The device of any one of embodiments A1-A53, wherein the device comprises a substrate.

[0143] A55: The device of embodiments A54, wherein the strip is attached to the substrate.

[0144] A56: The device of embodiments A54, wherein the substrate comprises a housing.

[0145] A57: The device of embodiments A54, wherein the substrate comprises a liquid impervious or non-porous surface.

[0146] A58: The device of embodiments A54, wherein the substrate comprises a flow control channel comprising the strip.

[0147] A59: The device of any one of embodiments A1-A53, having a limit of detection for the target analyte in a sample of at least 5 ng / mL.

[0148] A60: The device of embodiment A59, wherein the sample is selected from urine, blood or serum.

[0149] A61: The device of any one of embodiments A1-A60, further comprising a second detection region disposed on the strip, or disposed on the at least one porous membrane.

[0150] A62: The device of embodiment A61, wherein the second detection region is distal to the first detection region.

[0151] A63: The device of embodiment A61 or A62, wherein the second detection region comprises a binding agent that specifically binds to the first analyte-specific binding agent.

[0152] Bl: A lateral flow device for detecting a target analyte in a sample, the device comprising: a strip having a proximal end and a distal end, the strip comprising: (i) a samplereceiving region; (ii) at least one porous membrane; (iii) a flow control region comprising polyvinyl alcohol and mannitol; (iv) a filter region comprising a non-target antigen immobilized on the at least one porous membrane; and (v) a first detection region comprising a second analyte-specific binding agent immobilized on the at least one porous membrane.

[0153] B2. The device of embodiment Bl, further comprising colloidal gold nanoparticles comprising a polydopamine coating and a first analyte-specific binding agent.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT

[0154] B3. The device of embodiment Bl or B2, wherein the target analyte is fentanyl and the non-target antigen is an amphetamine.

[0155] B4. The device of any one of embodiments B 1 to B3, further comprising a buffering region.

[0156] Cl: A method for detecting a target analyte in a sample, the method comprising: (a) providing the lateral flow device of any embodiments A1-B4; (b) contacting a sample with the sample-receiving region; and (c) determining the presence, absence, or amount of detectable particles in the first detection region.

[0157] C2: The device of embodiment Cl, wherein the sample is contacted with the detectable particles prior to (b).

[0158] C3: The device of embodiment Cl or C2, further comprising determining the presence, absence, or amount of the gold nanoparticles in the second detection region.

[0159] C4: The device of any one of embodiments C1-C3, wherein the presence of gold nanoparticles in the first detection region indicates the presence of the target drug in the sample.

[0160] DI: A lateral flow device comprising: a flow control region located between a sample-receiving region and a detection region, the flow control region comprising water-soluble polymer and a sugar alcohol.

[0161] D2: The device of embodiment DI, wherein the water-soluble polymer comprises polyvinyl alcohol and the sugar alcohol comprises mannitol.

[0162] D3: The device of embodiment DI or D2, further comprising a filter region comprising a non-target antigen attached to an anchor moiety.

[0163] All patents, publications and abstracts cited above are incorporated herein by reference in their entireties. Various embodiments of the invention have been described in fulfillment of the various objectives of the invention. It should be recognized that these embodiments are merely illustrative of the principles of the present invention. Numerous modifications and adaptions thereof will be readily apparent to those skilled in the art without departing from the spirit and scope of the present invention as defined in the following claims.

[0164] In the embodiments described herein, all open-ended transitional phrases such as “comprising,” “including,” and / or “comprises,” and the like, can be used interchangeably with closed or semi-closed transitional phrases such as “consisting of’ and “consisting essentially of,” respectively. Accordingly, the embodiments disclosed herein may be practiced in the absence of any element which is not specifically disclosed herein. Thus, for example, in eachAttorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT instance herein the term “comprising” can be replaced with “consisting essentially of or “consisting of.’

[0165] As used herein, the meaning of “a,” “an,” or “the” includes singular and plural references unless the context clearly dictates otherwise.

[0166] All ranges disclosed herein are to be understood to encompass any and all subranges therein. For example, a stated range of “1 to 10” should be considered to include any and all subranges between (and inclusive of) the minimum value of 1 and the maximum value of 10; that is, all subranges beginning with a minimum value of 1 or more, e.g., 1 to 6.1, and ending with a maximum value of 10 or less, e.g., 5.5 to 10.

Claims

Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT WHAT IS CLAIMED IS:

1. A lateral flow device for detecting a target analyte in a sample, the device comprising: a strip having a proximal end and a distal end, the strip comprising:(i) a sample-receiving region disposed adjacent the proximal end;(ii) at least one porous membrane;(iii) a flow control region comprising a water-soluble polymer and / or a sugar alcohol; (iv) a filter region comprising a non-target antigen; and(v) a first detection region comprising a second analyte-specific binding agent.

2. The lateral flow device of claim 1, further comprising colloidal gold nanoparticles comprising a polydopamine coating and a first analyte-specific binding agent that binds specifically to the target analyte.

3. The lateral flow device of claim 2, wherein the colloidal gold nanoparticles are disposed on a particle region.

4. The lateral flow device of any one of claims 1 to 3, wherein the at least one porous membrane comprises or consists of nitrocellulose.

5. The lateral flow device of any one of claims 1 to 4, wherein the water-soluble polymer is selected from the group consisting of polyvinyl alcohol, polylactic acid, polyethylene glycol, alginate, and chitosan.

6. The lateral flow device of claim 5, wherein the water-soluble polymer comprises or consists of polyvinyl alcohol.

7. The lateral flow device of any one of claims 1 to 6, wherein the sugar alcohol comprises one or more of sorbitol, lactitol, mannitol, maltitol, erythritol, arabitol, ribitol, galactitol, and rhamnitol.

8. The lateral flow device of claim 7, wherein the sugar alcohol comprises or consists of mannitol.Attorney Docket No.: 057618-1549391Client Reference No.: LC-2024-17-WO-PCT9. The lateral flow device of any one claims 1 to 8, wherein the water-soluble polymer comprises or consists of polyvinyl alcohol and the sugar alcohol comprises or consists of mannitol.

10. The lateral flow device of any one of claims 1 to 9, wherein the non-target antigen is attached to an anchor moiety, and is irreversibly bound to the at least one porous membrane.

11. The lateral flow device of claim 10, wherein the anchor moiety comprises bovine serum albumin.

12. The lateral flow device of any one of claims 1 to 11, wherein the target analyte comprises or consists of fentanyl, or an analog, a derivative or metabolite thereof.

13. The lateral flow device of claim 12, wherein the metabolite of fentanyl comprises or consists of norfentanyl.

14. The lateral flow device of any one of claims 1 to 13, wherein the target analyte is different than the non-target antigen.

15. The lateral flow device of any one of claims 1 to 14, wherein the non-target antigen comprises or consists of a phenethylamine.

16. The lateral flow device of any one of claims 1 to 15, wherein the non-target antigen comprises or consists of an amphetamine, or an analog, a derivative or metabolite thereof.

17. A method for detecting a target drug in a sample, the method comprising:(a) providing the lateral flow device of any one of claims 1-16;(b) contacting a sample with the sample-receiving region of the lateral flow device; (c) determining a presence, absence, or amount of the gold nanoparticles in the first detection region, and(d) determining the presence, absence, or amount of the target drug in the sample according to the presence, absence, or amount of the gold nanoparticles determined in (c).

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