Witt Nina, Rodger Gillian, Vandesompele Jo, Benes Vladimir, Zumla Alimuddin, Rook Graham A, Huggett Jim F
Centre for Infectious Diseases and International Health, Windeyer Institute for Medical Sciences, University College London, London, United Kingdom.
J Biomol Tech. 2009 Dec;20(5):236-40.
Sensitive molecular methods, such as the PCR, can detect low-level contamination, and careful technique is required to reduce the impact of contaminants. Yet, some assays that are designed to detect high copy-number target sequences appear to be impossible to perform without contamination, and frequently, personnel or laboratory environment are held responsible as the source. This complicates diagnostic and research analysis when using molecular methods. To investigate the air specifically as a source of contamination, which might occur during PCR setup, we exposed tubes of water to the air of a laboratory and clean hood for up to 24 h. To increase the chances of contamination, we also investigated a busy open-plan office in the same way. All of the experiments showed the presence of human and rodent DNA contamination. However, there was no accumulation of the contamination in any of the environments investigated, suggesting that the air was not the source of contamination. Even the air from a busy open-plan office was a poor source of contamination for all of the DNA sequences investigated (human, bacterial, fungal, and rodent). This demonstrates that the personnel and immediate laboratory environment are not necessarily to blame for the observed contamination.
诸如聚合酶链反应(PCR)等灵敏的分子方法能够检测出低水平污染,并且需要采用精细的技术来降低污染物的影响。然而,一些旨在检测高拷贝数目标序列的检测方法似乎在无污染的情况下就无法进行,而且通常认为人员或实验室环境是污染源。这使得在使用分子方法时诊断和研究分析变得复杂。为了专门研究空气作为可能在PCR设置过程中出现的污染源,我们将装有水的试管暴露在实验室和洁净台的空气中长达24小时。为了增加污染的可能性,我们还以同样的方式研究了一个繁忙的开放式办公室。所有实验均显示存在人类和啮齿动物DNA污染。然而,在所研究的任何环境中污染都没有累积,这表明空气不是污染源。即使是来自繁忙开放式办公室的空气,对于所有所研究的DNA序列(人类、细菌、真菌和啮齿动物)来说也是较差的污染源。这表明观察到的污染不一定归咎于人员和实验室的直接环境。