Multianalyte molecular analysis using application-specific random particle arrays

Abstract

The present invention provides methods and apparatus for the application of a particle array in bioassay format to perform qualitative and/or quantitative molecular interaction analysis between two classes of molecules (an analyte and a binding agent). The methods and apparatus disclosed herein permit the determination of the presence or absence of association, the strength of association, and/or the rate of association and dissociation governing the binding interactions between the binding agents and the analyte molecules. The present invention is especially useful for performing multiplexed (parallel) assays for qualitative and/or quantitative analysis of binding interactions of a number of analyte molecules in a sample.

1 - 40 . (canceled)  
     
     
         41 . A method of real time monitoring of a multiplicity of target-oligonucleotide probe binding reactions carried out in the same reaction vessel, where detection of reaction results is performed by imaging signals which indicate binding of said targets to oligonucleotide probes, said oligonucleotide probes being bound to encoded carriers, wherein different types of oligonucleotide probes are bound to differently-encoded carriers and signals associated with differently-encoded carriers can be discriminated, comprising: 
 conjugating said carriers with oligonucleotide probes which include two complementary subsequences, separated by a third subsequence, and wherein a first complementary subsequence has a greater number of nucleotides which are complementary to the target subsequence than to the other complementary subsequence;    providing conditions such that said two complementary subsequences are annealed in the absence of target, but wherein the binding of the target subsequence to the first complementary subsequence causes separation of said two complementary subsequences and generation of a fluorescent signal;    forming said carriers into a planar array by placing them on a planar surface which is placed inside the reaction vessel;    adding targets, in solution, to the reaction vessel;    imaging said planar array to detect said signals at a first time and at least one other time, to thereby monitor difference in signals between the two times and to thus monitor the relative quantity target-ligand binding at said first and other times; and    identifying the types of ligands binding to targets by decoding the carriers associated with signals.    
     
     
         42 . The method of  claim 41  wherein the signals are monitored continuously.  
     
     
         43 . The method of  claim 41  wherein the images are recorded.  
     
     
         44 . The method of  claim 43  wherein binding of the target subsequence is detected by means of detecting fluorescence from a pair of fluorescent moieties respectively associated with the first and other complementary subsequence.  
     
     
         45 . The method of  claim 44  wherein the binding of the target subsequence is detected by means of detecting fluorescence from the donor in a donor-acceptor pair of fluorescent moieties respectively associated with the first and second subsequences (the members of the pair capable of interacting, when the first and second subsequences are annealed) by fluorescence energy transfer.