Absolute PCR Quantification

Abstract

The present application provides methods and devices for absolute quantification of polymerase chain reaction target nucleic acids. In particular, the methods and devices of the present application provide for splitting a nucleic acid sample to be analyzed into small, isolated volumes, conducting the method of polymerase chain reaction (PCR) on said volumes, detecting PCR amplification products, analyzing said detected PCR amplification products, performing absolute quantification of the PCR target and presenting said quantification results.

1 - 39 . (canceled) 
     
     
         40 . A method of sampling a target nucleic acid, comprising:
 a) providing a sample tray comprising a plurality of isolated reaction zones, wherein each isolated reaction zone contains a nucleic acid binding agent having a specific nucleic acid binding rate or specific nucleic acid binding capacity, wherein either the binding rates of the binding agents in different isolated reaction zones are variable binding rates, or the binding capacities of the binding agents in different isolated reaction zones are variable binding capacities;   b) exposing the sample tray to a sample comprising target nucleic acid under conditions wherein each isolated reaction is exposed to said sample for essentially the same amount of time;   c) removing unbound target nucleic acid from said sample tray;   d) detecting the target nucleic acid in said isolated reaction zones.   
     
     
         41 . The method of  claim 40 , wherein said variable binding rates are serial binding rates. 
     
     
         42 . The method of  claim 40 , wherein said nucleic acid binding agent comprises a gel matrix. 
     
     
         43 . The method of  claim 42 , wherein the specific nucleic acid binding rate of said gel matrix is determined by the rate at which said nucleic acid diffuses through said gel matrix. 
     
     
         44 . The method of  claim 42 , wherein the gels matrices in said different isolated reaction zones have differing gel porosities. 
     
     
         45 . The method of  claim 42 , wherein the gels matrices in said different isolated reaction zones have differing gel volumes. 
     
     
         46 . The method of  claim 41 , wherein the serial binding rates differ between isolated reaction zones by factors of 2. 
     
     
         47 - 49 . (canceled) 
     
     
         50 . The method of  claim 41 , wherein the serial binding rates differ between isolated reaction zones by factors of 1000. 
     
     
         51 . (canceled) 
     
     
         52 . The method of  claim 41 , wherein said variable binding capacities are serial binding capacities. 
     
     
         53 . The method of  claim 52 , wherein the serial binding capacities differ between isolated reaction zones by factors of 2. 
     
     
         54 - 56 . (canceled) 
     
     
         57 . The method of  claim 52 , wherein the serial binding capacities differ between isolated reaction zones by factors of 1000. 
     
     
         58 - 74 . (canceled)