# US 2018/0024129 A1 · Lateral flow device and method of use

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- Number: US 2018/0024129 A1
- Document ID: US20180024129A1
- Status: Published application
- Kind: A1
- Application number: 15/656,780
- Filed: 2017-07-21
- Published: 2018-01-25
- Earliest claimed: 2016-07-25
- Assignees: Bio-Rad Laboratories, Inc.
- Applicants: Bio-Rad Laboratories, Inc.
- Inventors: William Strong
- Web page: https://patentgrep.com/patent/US20180024129A1

## CPC

- G01N 33/54388: {based on lateral flow}
- C12Y 102/01012: Glyceraldehyde-3-phosphate dehydrogenase (phosphorylating) (1.2.1.12)
- C12Y 204/0203: NAD+ ADP-ribosyltransferase (2.4.2.30), i.e. tankyrase or poly(ADP-ribose) polymerase
- C12Y 207/11026: Tau-protein kinase (2.7.11.26)
- C12Y 306/05002: Small monomeric GTPase (3.6.5.2)
- G01N 33/525: {Multi-layer analytical elements}
- G01N 2333/47: Assays involving proteins of known structure or function as defined in the subgroups
- G01N 2333/90203: acting on the aldehyde or oxo group of donors (1.2)
- G01N 2333/91142: Pentosyltransferases (2.4.2)
- G01N 2333/912: transferring phosphorus containing groups, e.g. kinases (2.7)
- G01N 2333/914: Hydrolases (3)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- C12Y: ENZYMES

## Abstract

Lateral flow devices, methods and kits for performing lateral flow western blot assays are provided.

## Claims

### Claim 1

A lateral flow device comprising:
a wicking pad composed of a porous material, the wicking pad having a region for applying a substrate comprising immobilized analytes; and
wherein the wicking pad has a first end, a second end and two lateral edges;
a first reservoir comprising a stack of a plurality of reagent layers located on the first end of the wicking pad; and
a second reservoir comprising an absorbent pad located on the second end of the wicking pad.

### Claim 2 (depends on 1)

The device of claim 1, wherein each of the plurality of reagent layers comprises a reagent immobilized in an absorbent pad.

### Claim 3 (depends on 2)

The device of claim 2, wherein each of the plurality of reagent layers has a different reagent therein.

### Claim 4 (depends on 2)

The device of claim 2, wherein the reagent is selected from the group consisting of a primary antibody, a secondary antibody, a first wash solution, and a second wash solution.

### Claim 5 (depends on 1)

The device of claim 1, wherein the plurality of reagent layers, starting at a reagent layer in contact with the wicking pad, comprises a first reagent layer having a primary antibody, a second reagent layer having a first wash solution, a third reagent layer having a secondary antibody, and a fourth reagent layer having a second wash solution.

### Claim 6 (depends on 5)

The device of claim 5, wherein the fourth reagent layer is at least twice the thickness of the third reagent layer.

### Claim 7 (depends on 5)

The device of claim 5, further comprising a fifth reagent layer comprising the second wash solution.

### Claim 8 (depends on 5)

The device of claim 5, wherein the volume of the second wash solution is at least twice the volume of the secondary antibody.

### Claim 9 (depends on 5)

The device of claim 5, wherein each of the plurality of reagent layers is formed of an absorbent pad and at least a portion of the first reagent layer is in intimate contact with the wicking pad.

### Claim 10 (depends on 1)

The device of claim 1, wherein the analytes are proteins.

### Claim 11 (depends on 1)

The device of claim 1, wherein the device is sealed in a plastic casing.

### Claim 12 (depends on 11)

The device of claim 11, wherein the plastic casing comprises a molded bottom portion and a planar cover sealed to the bottom portion.

### Claim 13 (depends on 1)

The device of claim 1, wherein the wicking pad and reservoirs are dry.

### Claim 14 (depends on 1)

The device of claim 1, wherein at least one of the wicking pad and the first reservoir is wet.

### Claim 15 (depends on 1)

The device of claim 1, wherein the reagent layers are each formed of at least one absorbent material selected from the group consisting of cotton, glass fiber, cellulose, a cellulose fiber derivative, sintered glass, sintered polymer, sintered metal, and a synthetic polymer.

### Claim 16 (depends on 15)

The device of claim 15, wherein the synthetic polymer is selected from the group consisting of polyacrylamide, nylon, polypropylene, polyethylene, polystyrene, divinylbenzene, polyvinyl, polyvinyl difluoride, high density polyvinyl difluoride, a (C2-C6) monoolefin polymer, a vinylaromatic polymer, a vinylaminoaromatic polymer, a vinylhalide polymer, a (C1-C6) alkyl (meth)acrylate polymer, a(meth)acrylamide polymer, a vinyl pyrrolidone polymer, a vinyl pyridine polymer, a (C1-C6) hydroxyalkyl (meth)acrylate polymer, a (meth)acrylic acid polymer, an acrylamidomethylpropylsulfonic acid polymer, an N-hydroxy-containing (C1-C6) alkyl(meth)acrylamide polymer, and acrylonitrile.

### Claim 17 (depends on 1)

The device of claim 1, wherein each of the reagent layers further comprises a barrier layer on a portion of a lower surface.

### Claim 18 (depends on 1)

A kit for lateral flow, the kit comprising,
the device of claim 1.

### Claim 19 (depends on 1)

A method of performing a lateral flow assay, the method comprising:
providing the device of claims 1;
applying a running buffer to the wicking pad;
applying a substrate comprising proteins to the region for applying the substrate comprising analytes;
optionally wetting the first reservoir with the running buffer; and
allowing lateral flow of the running buffer from the first reservoir to the second reservoir such that reagents in the plurality of reagent layers are sequentially transported in the wicking pad and are contacted to the proteins on the substrate.

### Claim 20 (depends on 19)

The method of claim 19, wherein the allowing lateral flow step comprises allowing primary antibodies from a first reagent layer to bind to their target proteins, if present, on the substrate, followed by allowing a first wash solution from a second reagent layer to remove unbound primary antibodies from the substrate.

### Claim 21 (depends on 20)

The method of claim 20, wherein the allowing lateral flow step further comprises allowing secondary antibodies or a secondary detection reagent from a third reagent layer to contact the primary antibodies bound to their target proteins, if present, on the substrate.

### Claim 22 (depends on 21)

The method of claim 21, wherein the allowing lateral flow step further comprises allowing a second wash solution from a fourth reagent layer to remove unbound secondary antibodies from the substrate.

### Claim 23 (depends on 19)

The method of claim 19, further comprising applying a substantially uniform pressure to the first reservoir and the second reservoir.

### Claim 24 (depends on 19)

The method of claim 19, further comprising following binding of the primary antibodies to the target proteins, if present, (and optionally contact of secondary antibodies or secondary detection reagents to the primary antibodies), removing the substrate, and detecting the binding of the primary antibodies to the target proteins if present.

### Claim 25 (depends on 19)

The method of claim 19, wherein the allowing lateral flow step comprises allowing the reagents to follow a sinuous path as they flow through the first reservoir and into the wicking pad.