In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue.
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.
Strappe
P M
PM
Department of Pathology, University of Edinburgh, Western General Hospital, UK.
Wang
T H
TH
McKenzie
C A
CA
Lowrie
S
S
Simmonds
P
P
Bell
J E
JE
D001921:Brain / Q000821:virology*; D004279:DNA, Viral / Q000302:isolation & purification*; D005557:Formaldehyde; D015497:HIV-1 / Q000302:isolation & purification*; D006801:Humans; D008872:Microwaves; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D016707:Tissue Fixation
D004279:DNA, Viral; D005557:Formaldehyde