Methods of detecting targets on an arrary

Abstract

The invention relates to methods of detecting a target analyte in a biological sample using composite microsphere arrays having first and second assay locations. Preferred target analytes include nucleic acid, and more specifically, nucleic acid having one or more single nucleotide polymorphisms (SNPs).

What is claimed is:  
     
         1 . A method of detecting a target analyte in a biological sample, comprising: 
 a) providing a composite array comprising: 
 a substrate having a surface;  
 a first and a second assay location on said surface, wherein said assay locations comprise a population of microspheres, and wherein said microspheres comprise bioactive agents;  
 a physical partition separating said first assay location from said second assay location;  
   b) adding said biological sample to said first assay location under conditions sufficient to allow said target analyte to bind to said bioactive agents; and    c) detecting the binding of said bioactive agents to said target analyte.    
     
     
         2 . The method of  claim 1 , wherein the binding of said bioactive agents to said target analyte is detected by a change in an optical signature of the microspheres.  
     
     
         3 . The method of  claim 1 , wherein said target analyte comprises a nucleic acid.  
     
     
         4 . The method of  claim 3 , wherein said bioactive agents comprise a nucleic acid.  
     
     
         5 . The method of  claim 4 , wherein said microspheres further comprise labeled single-stranded nucleic acid sequences hybridized to said bioactive agents.  
     
     
         6 . The method of  claim 1 , wherein said microspheres are randomly distributed on said surface at said assay locations.  
     
     
         7 . The method of  claim 1 , wherein said microspheres are separated by a distance of less than 25 μm.  
     
     
         8 . The method of  claim 1 , wherein said microspheres are separated by a distance of less than 15 μm.  
     
     
         9 . The method of  claim 1 , further comprising detecting a target analyte in a second biological sample by adding said second biological sample to said second assay location and thereafter detecting the binding of said bioactive agents to said target analyte.  
     
     
         10 . The method of  claim 1 , wherein said substrate comprises a microscope slide.  
     
     
         11 . The method of  claim 1 , wherein said first and second assay locations are separately sealed to form hybridization chambers.  
     
     
         12 . The method of  claim 11 , wherein said hybridization chambers comprise flexible membranes.  
     
     
         13 . The method of  claim 1 , wherein said physical partition is a non-permanent sealant.  
     
     
         14 . The method of  claim 13 , wherein said non-permanent sealant is selected from the group consisting of: rubber, silicon, petroleum jelly, wax and parafilm.  
     
     
         15 . The method of  claim 1 , wherein said physical partition is a ridge or rim of sufficient width and height to prevent said bioactive agents from moving from said first assay location to said second assay location.  
     
     
         16 . The method of  claim 1 , wherein said physical partition is a trough of sufficient width and depth to prevent said bioactive agents from moving from said first assay location to said second assay location.  
     
     
         17 . The method of  claim 1 , wherein said physical partition is a gasket.  
     
     
         18 . The method of  claim 17 , wherein said gasket is adapted to fit within an indentation or channel on the substrate.  
     
     
         19 . The method of  claim 1 , wherein said target analyte comprises a detectable optical label.  
     
     
         20 . The method of  claim 19 , wherein said optical label is a fluorophore.  
     
     
         21 . The method of  claim 1 , wherein said biological sample comprises blood.  
     
     
         22 . The method of  claim 1 , wherein said bioactive agents comprise a detectable optical label.  
     
     
         23 . The method of  claim 1 , wherein said detectable label is a fluorophore.  
     
     
         24 . A method of detecting a target nucleic acid in a biological sample, wherein said target nucleic acid comprises one or more single nucleotide polymorphisms (SNPs) at one or more predetermined positions, comprising: 
 a) providing a composite array comprising: 
 a substrate having a surface;  
 a population of microspheres, wherein said microspheres are linked to capture probes configured to bind to said target nucleic acid at said one or more predetermined positions;  
 a first and second assay location on said surface, wherein said assay locations comprise said population of microspheres;  
 a physical partition separating said first assay location from said second assay location;  
   b) adding said biological sample to said first assay location under conditions sufficient to allow said target nucleic acid to bind to said capture probes; and    c) detecting the binding of said target nucleic acid to said capture probes.    
     
     
         25 . The method of  claim 24 , wherein the binding of said capture probes to said target nucleic acid is detected by a change in an optical signature of the microspheres.  
     
     
         26 . The method of  claim 24 , wherein said microspheres are randomly distributed on said surface at said assay locations.  
     
     
         27 . The method of  claim 24 , wherein said microspheres are separated by a distance of less than 25 μm.  
     
     
         28 . The method of  claim 24 , wherein said microspheres are separated by a distance of less than 15 μm.  
     
     
         29 . The method of  claim 24 , further comprising detecting a target analyte in a second biological sample by adding said second biological sample to said second assay location and thereafter detecting the binding of said capture probes to said target analyte.  
     
     
         30 . The method of  claim 24 , wherein said substrate comprises a microscope slide.  
     
     
         31 . The method of  claim 24 , further comprising adding a SNP allele-specific probe, configured to hybridize to said target nucleic acid, wherein said SNP allele-specific probe is labeled with a fluorophore.  
     
     
         32 . The method of  claim 24 , wherein said physical partition is a non-permanent sealant.  
     
     
         33 . The method of  claim 32 , wherein said non-permanent sealant is selected from the group consisting of rubber, silicon, petroleum jelly, wax and parafilm.  
     
     
         34 . The method of  claim 24 , wherein said physical partition is a ridge or rim of sufficient width and height to prevent said capture probes from moving from said first assay location to said second assay location.  
     
     
         35 . The method of  claim 24 , wherein said physical partition is a trough of sufficient width and depth to prevent said capture probes from moving from said first assay location to said second assay location.  
     
     
         36 . The method of  claim 24 , wherein said physical partition is a gasket.  
     
     
         37 . The method of  claim 36 , wherein said gasket is adapted to fit within an indentation or channel on the substrate.  
     
     
         38 . The method of  claim 24 , wherein said biological sample comprises blood.  
     
     
         39 . The method of  claim 24 , wherein said target nucleic acid having one or more SNPs is causatively linked to a human disease.  
     
     
         40 . The method of  claim 39 , wherein said disease is Alzheimer's disease.  
     
     
         41 . The method of  claim 24 , wherein said target nucleic acid comprises a detectable optical label.  
     
     
         42 . The method of  claim 41 , wherein said optical label is a fluorophore.  
     
     
         43 . The method of  claim 24 , wherein said microspheres further comprise labeled single-stranded nucleic acid sequences hybridized to said capture probes.  
     
     
         44 . The method of  claim 24 , wherein said capture probes comprise a detectable optical label.  
     
     
         45 . The method of  claim 44 , wherein said optical label is a fluorophore.