Method of Performing Digital PCR

Abstract

A method of detection of a target nucleic acid is provided. The method includes fractionating a sample into a plurality of sample volumes wherein more than 50% of the fractions contain no more than 1 target nucleic acid molecule per sample volumes, and subjecting the plurality of sample volumes to conditions for amplification. The method further includes detecting a change in ion concentration in a sample volume wherein a target nucleic acid is present, counting the number of fractions with an amplified target nucleic acid, and determining the quantity of target nucleic acid in the sample.

What is claimed is: 
     
         1 . A device for performing PCR comprising:
 a sample holder configured to fractionate a sample into a plurality of sample volumes wherein more than 50% of the fractions contain no more than 1 target nucleic molecules per sample volume and further configured to hold a sample;   a sensing layer in chemical communication with the sample holding device; and   at least one sensor in electrical communication with the sensing layer.   
     
     
         2 . The device of  claim 1 , wherein the sample holding device is a matrix. 
     
     
         3 . The device of  claim 2 , wherein the matrix is an extracellular matrix. 
     
     
         4 . The device of  claim 1 , wherein the sample holding device is a gel. 
     
     
         5 . The device of  claim 4 , wherein the gel is a hydrogel. 
     
     
         6 . The device of  claim 1 , wherein the sample holding device is located around at least a portion of the periphery of the sensor. 
     
     
         7 . A device for performing PCR comprising:
 a sample holder configured hold a sample and to fractionate the sample into a plurality of sample volumes, wherein the sample contains an initial quantity of a target nucleic acid and wherein more than 50% of the fractions contain no more than 1 target nucleic molecule of the target nucleic acid per sample volume;   a thermal cycler configured to amplify or enrich the sample to produce an amplified quantity of the target nucleic acid molecule so that an ion concentration in one or more of the sample volumes is changed; and   an ion detector comprising a plurality of sensors in communication with the sample volumes, the ion detector configured to detect a change in ion concentration in the one or more of sample volumes;   wherein the system is configured to determine the initial quantity of the target nucleic acid from the detected change(s) in the one or more sample volumes.   
     
     
         8 . The device of  claim 7 , wherein the sample holding device is a matrix. 
     
     
         9 . The device of  claim 8 , wherein the matrix is an extracellular matrix. 
     
     
         10 . The device of  claim 7 , wherein the sample holding device is a gel. 
     
     
         11 . The device of  claim 10 , wherein the gel is a hydrogel. 
     
     
         12 . The device of  claim 7 , wherein the sample holding device is located around at least a portion of the periphery of the sensor. 
     
     
         13 . A device for performing PCR comprising:
 a sample holder configured hold a sample and to fractionate the sample into a plurality of sample volumes, wherein more than 50% of the fractions contain no more than 1 target nucleic molecules per sample volume;   a thermal cycler configured to amplify or enrich the sample to produce an amplified quantity of the target nucleic acid molecule so that an ion concentration in one or more of the sample volumes is changed;   a sensing layer in chemical communication with the sample holding device; and   at least one sensor in electrical communication with the sensing layer;   wherein the system is configured to determine the initial quantity of the target nucleic acid from ion concentration change(s) sensed in the at least one sensor.   
     
     
         14 . The device of  claim 13 , wherein the sample holding device is a matrix. 
     
     
         15 . The device of  claim 14 , wherein the matrix is an extracellular matrix. 
     
     
         16 . The device of  claim 13 , wherein the sample holding device is a gel. 
     
     
         17 . The device of  claim 16 , wherein the gel is a hydrogel. 
     
     
         18 . The device of  claim 13 , wherein the sample holding device is located around at least a portion of the periphery of the sensor.