Grote Hans Jürgen, Schmiemann Viola, Sarbia Mario, Böcking Alfred
Institute of Cytopathology, Heinrich-Heine-University, Moorenstrasse 5, D-40225 Düsseldorf, Germany.
Anal Cell Pathol. 2003;25(2):83-8. doi: 10.1155/2003/354796.
To date, there are only few systematic reports on the quality of DNA extracted from routine diagnostic cytologic specimens. It was the aim of the present study to evaluate the ability of 50% ethanol/2% carbowax (Saccomanno fixative) to preserve bronchial secretions with high quality genomic DNA as well as to compare different DNA extraction methods.
DNA was extracted from 45 bronchial aspirates by four different extraction protocols. Beside DNA yield, DNA quality with regard to purity, integrity, and PCR success rate were investigated.
No fragmentation of sample DNA due to the fixative was detected. It was preserved as high molecular weight DNA. DNA yield, purity, and integrity were dependent on the DNA extraction method to some extend. Irrespective of the DNA extraction method the PCR success rate for amplification of beta-globin gene fragments (268, 536, and 989 bp) was 100%.
A fixative containing 50% ethanol/2% carbowax preserves high quality DNA which is well suited for PCR-based assays regardless of the extraction protocol used. The selection of the DNA extraction protocol has to be adjusted to the circumstances of application.
迄今为止，关于从常规诊断性细胞学标本中提取的DNA质量的系统性报告很少。本研究的目的是评估50%乙醇/2%卡波蜡（萨科曼诺固定液）保存具有高质量基因组DNA的支气管分泌物的能力，并比较不同的DNA提取方法。
通过四种不同的提取方案从45份支气管吸出物中提取DNA。除了DNA产量外，还研究了DNA在纯度、完整性和PCR成功率方面的质量。
未检测到由于固定液导致的样本DNA片段化。它被保存为高分子量DNA。DNA产量、纯度和完整性在一定程度上取决于DNA提取方法。无论采用何种DNA提取方法，β-珠蛋白基因片段（268、536和989 bp）的PCR成功率均为100%。
含有50%乙醇/2%卡波蜡的固定液可保存高质量的DNA，无论使用何种提取方案，该DNA都非常适合基于PCR的检测。DNA提取方案的选择必须根据应用情况进行调整。
