Colocalization-by-linkage sandwich assays

Abstract

There are provided methods and systems for detecting and/or quantifying an analyte. In particular, there are provided methods and systems for simultaneous detection and/or quantitation of two or more analytes in a sample. In some embodiments, there are provided colocalization-by-linkage assays on microparticles (CLAMP) comprising two sets of binders pre-assembled on a support, such that the two sets of binders are colocalized before contacting the sample.

1 . A biomolecule complex for the detection or quantification of an analyte in a sample, comprising:
 (a) a support complex comprising:
 (i) a support; 
 (ii) a capture reagent coupled with the support; 
 (iii) an anchor element coupled with the support; 
 (iv) a detection reagent coupled with a hook element, wherein the hook element is releasably coupled with the anchor element; and 
   (b) a detectably-labeled displacer reagent capable of coupling with the hook element and decoupling the hook element from the anchor element;   
       wherein the capture reagent and the detection reagent are configured to simultaneously couple with the analyte. 
     
     
         2 . The biomolecule complex of  claim 1 , wherein the support complex further comprises the detectably-labeled displacer reagent when (i) the detectably-labeled displacer reagent is coupled to the hook element, and (ii) the analyte is coupled to the capture reagent and the detection reagent. 
     
     
         3 . The biomolecule complex of  claim 1 , further wherein the support complex does not comprise the detectably-labeled displacer reagent when (i) the detectably-labeled displacer reagent is coupled to the hook element, and (ii) the analyte is not coupled to one or both of the capture reagent and detection reagent. 
     
     
         4 . The biomolecule complex of  claim 1 , wherein the anchor element and the hook element are an oligonucleotide. 
     
     
         5 . The biomolecule complex of  claim 4 , wherein the detectably-labeled displacer reagent comprises an oligonucleotide. 
     
     
         6 . The biomolecule complex of  claim 5 , wherein the anchor element comprises an anchor sequence and the hook element comprises a linker sequence complementary to the anchor sequence. 
     
     
         7 . The biomolecule complex of  claim 6 , wherein the hook element comprises an additional sequence adjacent to the linker sequence and the detectably-labeled displacer reagent comprises a displacer sequence complementary to the additional sequence and at least a portion of the linker sequence. 
     
     
         8 . The biomolecule complex of  claim 7 , wherein the displacer sequence is complementary to the linker sequence. 
     
     
         9 . The biomolecule complex of  claim 7 , wherein the additional sequence comprises at least one nucleotide. 
     
     
         10 . The biomolecule complex of  claim 7 , wherein the displacer sequence has a melting temperature greater than that of the anchor sequence. 
     
     
         11 . The biomolecule complex of  claim 1 , wherein the sample is a biological sample. 
     
     
         12 . The biomolecule complex of  claim 11 , wherein the sample is a bodily fluid, a whole blood sample, a cell supernatant, an extract, a cell extract, a cell lysate, a tissue lysate, a solution comprising nucleic acid molecules, or a solution comprising proteins. 
     
     
         13 . The biomolecule complex of  claim 1 , wherein the capture reagent is selected from the group consisting of: an antibody or an antigen-binding fragment thereof, an aptamer, a modified aptamer, a somamer, an affimer, an antigen, a protein, a polypeptide, a multi-protein complex, an exosome, an oligonucleotide, a low molecular weight compound, and any combination thereof. 
     
     
         14 . The biomolecule complex of  claim 1 , wherein the detection reagent is selected from the group consisting of: an antibody or an antigen-binding fragment thereof, an aptamer, a modified aptamer, a somamer, an affimer, an antigen, a protein, a polypeptide, a multi-protein complex, an exosome, an oligonucleotide, a low molecular weight compound, and any combination thereof. 
     
     
         15 . The biomolecule complex of  claim 1 , wherein the capture reagent and the detection reagent are both antibodies or antigen-binding fragments thereof. 
     
     
         16 . The biomolecule complex of  claim 15 , wherein the capture reagent and the detection reagent are a different antibody or antigen-binding fragment thereof and bind to a different epitope on the analyte. 
     
     
         17 . The biomolecule complex of  claim 1 , wherein the detectably-labeled displacer reagent comprises a detectable label selected from the group consisting of a fluorescent polymer, a biotin molecule, a fluorophore, an enzyme, a nucleic acid enzyme, a riboswitch, an enzyme substrate, a specific nucleic acid sequence, and any combination thereof. 
     
     
         18 . The biomolecule complex of  claim 1 , wherein the support is a microparticle, a nanoparticle, a well in a plate, an array, a microfluidic chip, a lateral flow strip, a slide, a porous polymer, or a hydrogel. 
     
     
         19 . The biomolecule complex of  claim 1 , wherein the support complex comprises a plurality of the capture reagent, a plurality of the anchor element, and a plurality of the detection reagents coupled with the hook element. 
     
     
         20 . The biomolecule complex of  claim 1 , wherein the detection reagent is coupled with a single hook element. 
     
     
         21 - 24 . (canceled)