Comparative studies on the detection of hepatitis B virus DNA in frozen and paraffin sections by the polymerase chain reaction.
The polymerase chain reaction (PCR) is an extremely sensitive technique that has been used for detection of DNA sequences in formalin-fixed, paraffin-embedded tissues. In order to verify that hepatitis B virus (HBV) DNA sequences are adequately preserved in routinely processed liver tissues, we performed PCR with five different primer pairs for HBV sequences on DNA extracted by two different methods from paraffin and frozen liver sections. The amount of PCR products obtained with DNA templates extracted by the proteinase K-SDS method from frozen sections was significantly larger than that from paraffin sections. However, boiling of deparaffinized sections in water containing Chelex-100 resulted in ample amounts of PCR products irrespective of the primers used. On Southern blots, the location of the bands of amplified DNA obtained by the different methods was consistent with the predicted size, suggesting that the viral sequences had not been altered by processing. Although freezing of fresh tissue yields quantitatively more HBV DNA, formalin fixation qualitatively preserves the viral DNA sequences adequately for detection by PCR. Therefore, formalin-fixed, paraffin-embedded tissues may be used for the detection of viral DNA sequences by PCR. Application of the described procedure to routinely processed tissues significantly broadens the applicability of this powerful diagnostic and investigative method.
Chen
M L
ML
Department of Pathology, Tulane University School of Medicine, New Orleans, Louisiana.
Shieh
Y S
YS
Shim
K S
KS
Gerber
M A
MA
D001483:Base Sequence; D001706:Biopsy; D015139:Blotting, Southern; D016616:Cryoultramicrotomy; D004279:DNA, Viral / Q000032:analysis* / Q000235:genetics; D006515:Hepatitis B virus / Q000235:genetics* / Q000302:isolation & purification; D006651:Histocytochemistry; D006801:Humans; D008099:Liver / Q000737:chemistry / Q000382:microbiology / Q000473:pathology; D008969:Molecular Sequence Data; D010232:Paraffin; D016133:Polymerase Chain Reaction
D004279:DNA, Viral; D010232:Paraffin