Systems and methods for detection of analytes in biological fluids

Abstract

The invention provides a heterogeneous immunoassay for detection of antibodies and antigens based on specific antigen-antibody immune complex formation with multiple antigen-bearing conjugate components. The invention further provides means for optimizing the assay format for the detection of both low and high-affinity antibodies, and provides means for quantitative detection of both antibody and the corresponding antigen present in a sample.

1 . A method for detecting an antibody that binds to an antigen or hapten in a sample comprising: 
 providing a first component comprising a ligand linked to a solid phase adsorbing composition though a chemical spacer;    providing a second component comprising a ligand-binding moiety and the antigen or hapten that binds the antibody;    providing a third component comprising an antigen or hapten and a label;    contacting the first component with a solid phase so as to immobilize the first component;    contacting the antibody sample with the first, second and third components and allowing the third component to form an immune complex with the first and second components by binding to the second component through the antibody present in the antibody sample; and    separating uncomplexed third component from the solid phase and detecting the label present in the immune complex, wherein detection of said label indicates the presence of the antibody in the antibody sample,    thereby detecting the antibody in the sample.    
   
   
       2 . The method of  claim 1 , wherein the ligand is biotin.  
   
   
       3 . The method of  claim 1 , wherein the ligand-binding moiety is avidin or streptavidin.  
   
   
       4 . The method of  claim 1 , wherein the solid phase adsorbing composition is an albumin.  
   
   
       5 . (canceled)  
   
   
       6 . (canceled)  
   
   
       7 . (canceled)  
   
   
       8 . (canceled)  
   
   
       9 . (canceled)  
   
   
       10 . (canceled)  
   
   
       11 . (canceled)  
   
   
       12 . The method of  claim 1 , wherein the separation of uncomplexed third component label from the solid phase is achieved by washing unbound third component label from the solid phase.  
   
   
       13 . The method of  claim 1 , wherein the quantity of antigen or hapten in the third component conjugate is greater than the quantity of antigen or hapten in the second component conjugate.  
   
   
       14 . (canceled)  
   
   
       15 . (canceled)  
   
   
       16 . (canceled)  
   
   
       17 . (canceled)  
   
   
       18 . (canceled)  
   
   
       19 . (canceled)  
   
   
       20 . (canceled)  
   
   
       21 . The method of  claim 1 , wherein the third component comprises an antigen or hapten conjugated indirectly to a label through a chemical spacer.  
   
   
       22 . (canceled)  
   
   
       23 . (canceled)  
   
   
       24 . (canceled)  
   
   
       25 . (canceled)  
   
   
       26 . (canceled)  
   
   
       27 . The method of  claim 1 , wherein the second and third component conjugates each comprise two or more different antigens or haptens.  
   
   
       28 . (canceled)  
   
   
       29 . The method of  claim 1 , wherein the antigens are peptides derived from VLSe protein (C6) and OspC protein (C10) of  B. burgdorferi  for the diagnosis of Lyme borreloiosis.  
   
   
       30 . The method of  claim 1 , further comprising quantitation of the antibody.  
   
   
       31 . The method of  claim 1 , further comprising contacting the mixture of the antibody sample with the first, second and third components with a sample containing the antigen or hapten for the quantitative detection of said antigen or hapten in said sample.  
   
   
       32 . A kit comprising the first, second and third components of  claim 1 .  
   
   
       33 . The kit of  claim 32 , further comprising an antibody standard.  
   
   
       34 . The kit of  claim 32 , wherein the second and third components are supplied as dry samples.  
   
   
       35 . The kit of  claim 32 , wherein the second and third components comprise more than one antigen or hapten molecule.  
   
   
       36 . A composition of matter comprising a ligand linked to a solid phase adsorbing composition though a chemical spacer.  
   
   
       37 . The composition of matter of  claim 36 , wherein the ligand is biotin.  
   
   
       38 . The composition of matter of  claim 36 , wherein the solid phase adsorbing composition is bovine serum albumin.  
   
   
       39 . The composition of matter of  claim 36 , wherein the chemical spacer is polyethylene glycol.  
   
   
       40 . The composition of matter of  claim 36 , wherein the ligand is biotin, the solid phase adsorbing composition is bovine serum albumin and the chemical spacer is polyethylene glycol.