Williams D L, Gillis T P, Dupree W G
Laboratory Research Branch, G. W. Long Hansen's Disease Center, Louisiana State University, Baton Rouge 70894, USA.
J Clin Microbiol. 1995 Jun;33(6):1558-61. doi: 10.1128/jcm.33.6.1558-1561.1995.
The effect of ethanol fixation on PCR detection and viability of Mycobacterium tuberculosis in human sputum sediments was evaluated. M. tuberculosis seeded into sputum sediments was efficiently killed when treated for 1 h with 50, 70, or 95% ethanol. PCR amplification of a 123-bp fragment of the M. tuberculosis-specific IS6110 was not affected in ethanol-treated samples even when fixation was extended to 24 h. Ethanol fixation of sputum sediments did not affect the PCR detection of M. tuberculosis in clinical samples. PCR results from ethanol-treated clinical samples containing M. tuberculosis (smear positive and smear negative) or other respiratory pathogens correlated directly with the results by conventional detection methods for M. tuberculosis. Our results show that ethanol fixation of human sputum sediments containing M. tuberculosis significantly reduces the potential exposure of workers to viable M. tuberculosis without affecting DNA analysis by PCR. Also, ethanol fixation of sputum sediments provides a simple and inexpensive way to store and transport clinical specimens identified for DNA-based diagnostics without refrigeration.
评估了乙醇固定对人痰液沉淀物中结核分枝杆菌PCR检测及活力的影响。接种于痰液沉淀物中的结核分枝杆菌用50%、70%或95%乙醇处理1小时后可被有效杀灭。即使固定时间延长至24小时，经乙醇处理的样本中结核分枝杆菌特异性IS6110的123bp片段的PCR扩增也不受影响。痰液沉淀物的乙醇固定不影响临床样本中结核分枝杆菌的PCR检测。含有结核分枝杆菌（涂片阳性和涂片阴性）或其他呼吸道病原体的乙醇处理临床样本的PCR结果与结核分枝杆菌的传统检测方法结果直接相关。我们的结果表明，对含结核分枝杆菌的人痰液沉淀物进行乙醇固定可显著降低工作人员接触活结核分枝杆菌的可能性，同时不影响PCR的DNA分析。此外，痰液沉淀物的乙醇固定为无需冷藏储存和运输用于基于DNA诊断的临床标本提供了一种简单且廉价的方法。
