Using 2-step PCR and restriction endonuclease digestion to detect K-ras mutation in paraffin-embedded tissues: is it reliable?
In this study, we detected the existence of both wild type and mutant K-ras DNA on the cutting board and in the solutions for tissue processing in our pathology laboratory. The detection rate of K-ras mutant DNA inversely correlated with the frequency of the solution changed, implying that DNA contaminant including K-ras mutant may be accumulated while increasing specimens were processed. The exact routes of contamination during tissue processing remain to be determined. Nevertheless, this study unveils the possibility of DNA contamination in the laboratory that can be amplified by PCR analysis, and precaution should be taken while we interpret our PCR data from paraffin-embedded specimens. The work also highlights the necessity of contamination control procedures, including specific laboratory construction, environmental control, and strict hygiene standards for laboratory equipment and personnel. Moreover, fresh specimens seem more reliable than archival materials for PCR diagnosis.
Su
W C
WC
Department of Internal Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan, ROC.
Chow
N H
NH
Huang
C H
CH
Liu
H S
HS
D004262:DNA Restriction Enzymes / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D011905:Genes, ras / Q000235:genetics*; D006801:Humans; D009154:Mutation / Q000235:genetics; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA; D014407:Tumor Cells, Cultured
D004262:DNA Restriction Enzymes; C046545:CCWGG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific