Signal amplification in lateral flow and related immunoassays
The present invention provides methods, devices, compositions (e.g., capture complexes), and kits useful for enhancing the detection of antibodies in a test sample. The methods, devices, and compositions utilize detectable Fc-binding molecules such as Protein A, Protein G, and/or an Fc-specific antibody to amplify the signal of a detected antibody in immunoassays, such as lateral flow assays.
1. A lateral flow assay device comprising
(a) a conjugate region comprising (i) a mobilizable first detector capable of binding to the Fc region of an antibody in a sample, wherein the first detector comprises a protein A and/or protein G, wherein the protein A and/or protein G is conjugated to a gold nanoparticle, and (ii) a mobilizable second detector, wherein the second detector comprises a first antigen or antigenic peptide conjugated to a gold nanoparticle, wherein said first antigen or antigenic peptide is capable of specifically binding to a variable region of the antibody in the sample; and
(b) a test region comprising an immobilized capture entity, wherein the capture entity comprises a second antigen or antigenic peptide, wherein said second antigen or antigenic peptide is capable of specifically binding to the variable region of the antibody in the sample.
2. The lateral flow assay device of claim 1 , wherein the device further comprises a control region comprising an antibody that binds to the first detector.
3. The lateral flow assay device of claim 2 , wherein the antibody that binds to the first detector is an anti-protein A or an anti-protein G antibody.
4. The lateral flow assay of claim 3 , wherein the anti-protein A antibody is a chicken anti-Protein A IgG antibody.
5. The lateral flow assay device of claim 2 , wherein the control region is located on a nitrocellulose membrane.
6. The lateral flow assay device of claim 2 , wherein the test and control regions are located on a nitrocellulose membrane, and wherein the test and control regions do not overlap.
7. The lateral flow assay device of claim 1 , wherein the test region is located on a nitrocellulose membrane.
8. The lateral flow assay device of claim 1 , wherein the conjugate and test regions are in fluid communication such that in operation a liquid sample comprising an antibody flows from the conjugate region to the test region.
9. The lateral flow assay device of claim 1 , wherein the device further comprises a sample loading region, wherein the conjugate region is positioned downstream of the sample loading region.
10. The lateral flow assay device of claim 1 , wherein the device further comprises an absorbent pad positioned downstream of the test region.
11. The lateral flow assay device of claim 1 , wherein said first antigen or antigenic peptide is from an organism selected from the group consisting of heartworm, Ehrlichia canis, Ehrlichia chaffeensis, Ehrlichia ewingii, Borrelia burgdorferi, Borrelia afzelii, Borrelia garinii, Anaplasma phagocytophilum, Anaplasma platys, feline leukemia virus, parvovirus, influenza A strain, influenza B strain, avian influenza virus, respiratory syncytial virus, Legionella , adenovirus, rotavirus, feline immunodeficiency virus, human immunodeficiency virus, and Group A Streptococcus.
12. The lateral flow assay device of claim 1 , wherein said second antigen or antigenic peptide is from an organism selected from the group consisting of heartworm, Ehrlichia canis, Ehrlichia chaffeensis, Ehrlichia ewingii, Borrelia burgdorferi, Borrelia afzelii, Borrelia garinii, Anaplasma phagocytophilum, Anaplasma platys , feline leukemia virus, parvovirus, influenza A strain, influenza B strain, avian influenza virus, respiratory syncytial virus, Legionella , adenovirus, rotavirus, feline immunodeficiency virus, human immunodeficiency virus, and Group A Streptococcus.
13. The lateral flow assay device of claim 1 , wherein the second detector and the capture entity comprise the same antigen or antigenic peptide.
14. The lateral flow assay device of claim 1 , wherein said first antigen or antigenic peptide and/or said second antigen or antigenic peptide is conjugated to bovine serum albumin (BSA).
15. A kit comprising the lateral flow assay device of claim 1 and instructions for using the device to detect an antibody in a test sample.
16. A method for determining the presence of an antibody in a sample, comprising contacting the test sample with the lateral flow assay device of claim 1 , and detecting the presence of a signal from the test region, wherein the presence of the signal is indicative of the presence of the antibody in the sample.