APPLICATION OF PCR FOR DETECTION AND IDENTIFICATION OF MYCOPLASMA CONTAMINATION IN VIRUS STOCKS

A nested polymerase chain reaction (PCR) was used to detect and identi fy mycoplasma contaminants in viral stocks. The results of the PCR ass ay proved to be a sensitive and accurate indicator of the true status of the stock tested. Those samples positive by agar culture or Hoechst stain were also positive by PCR. Those samples that were inconclusive by Hoechst stain (10.05%) could be clearly determined to be mycoplasm a positive or negative by PCR. The PCR assay also detected those fasti dious species of mycoplasma that gave false negative results by the di rect culture method. In many respects the PCR-based mycoplasma detecti on method described is superior to the agar culture and Hoechst staini ng detection methods. In this study, the PCR assay detected substantia lly more mycoplasma-positive viral stocks than did the agar culture as say. Due to its speed, sensitivity,and reliability, the PCR assay is o f particular value in monitoring the process of removing mycoplasma fr om contaminated stocks. Furthermore, the PCR amplification products ca n be analyzed by restriction analysis to rapidly identify the species of the mycoplasma contaminating the stock tested.