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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.

Type

Journal article

Journal

J Virol Methods

Publication Date

02/1998

Volume

70

Pages

119 - 127

Keywords

Brain, DNA, Viral, Formaldehyde, HIV-1, Humans, Microwaves, Paraffin Embedding, Polymerase Chain Reaction, Sensitivity and Specificity, Tissue Fixation