Patent ID: 7186507

Claim:
A method for in situ amplification and detection of a target nucleic acid sequence in a fixed, permeabilized cell comprising: (a) selecting a forward and a reverse primer for said target nucleic acid, wherein the forward and reverse primers flank at least one restriction enzyme site within said target nucleic acid sequence; (b) contacting the fixed, permeabilized cell with a restriction endonuclease composition comprising at least one restriction endonuclease that cuts a genomic nucleic acid sequence flanked by the forward and reverse primers specific for said target nucleic acid, to produce a restriction digest; (c) contacting said restriction digest in the cell with a DNAse to produce a DNAse digested cell; (d) incubating said cell with a reverse transcriptase (RT) cocktail comprising an RT enzyme and a RT primer specific for said target nucleic acid to produce a cDNA; and (e) amplifying said cDNA using a PCR reaction in the presence of the forward and reverse primers wherein at least one of said forward and reverse primers is labeled; and (f) detecting the amplified cDNA.