In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue

Abstract A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. Two methods are compared to detect the biotinylated amplified product, the use of an avidin–biotin–alkaline phosphatase complex (ABC) and the application of tyramide signal amplification (TSA) which allows both chromogenic and fluorescence detection. TSA detection enhances the sensitivity of IS-PCR, permitting fewer PCR cycles and preserving tissue morphology.