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 capture complex comprising a capture entity immobilized on a surface, an antibody, a first detector, and a second detector, wherein the capture entity binds to a variable region of the antibody and wherein the first detector comprises a Fc-binding molecule conjugated to a first detectable entity, wherein the Fc-binding molecule of the first detector is bound to the Fc region of the antibody, and wherein the second detector comprises an antigen or antigenic peptide specifically bound to a variable region of the antibody.
2. The capture complex of claim 1 , wherein the capture complex is immobilized on a test region of the surface.
3. The capture complex of claim 1 , wherein the second detector is conjugated to a second detectable entity.
4. The capture complex of claim 3 , wherein the first and second detectable entities are gold nanoparticles.
5. The capture complex of claim 3 , wherein the first detector and the second detector are present in a ratio of about 20:1 to about 1:1.
6. The capture complex of claim 1 , wherein the Fc-binding molecule is protein A and/or protein G.
7. The capture complex of claim 6 , wherein the Fc-binding molecule comprises protein A and protein G, and wherein protein A and protein G are present in a ratio of about 10:1 to about 1:10.
8. The capture complex of claim 1 , wherein the capture entity is an antigen or antigenic peptide.
9. The capture complex of claim 1 , wherein the wherein the capture entity is an antigen or antigenic peptide 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.
10. The capture complex of claim 1 , wherein the first detectable entity is a metal nanoparticle, metal nanoshell, fluorophore, or colored latex particle.
11. The capture complex of claim 10 , wherein the metallic nanoparticle or metallic nanoshell is selected from the group consisting of gold particles, silver particles, copper particles, platinum particles, cadmium particles, composite particles, gold hollow spheres, gold-coated silica nanoshells, and silica-coated gold shells.
12. The capture complex of claim 1 , wherein the surface is a flow path in a lateral flow assay device, a surface of a microtiter plate or a flow path in an analytical rotor.