Fan K, Tano H, Kitajima M, Tamatsukuri S, Furuichi Y, Hayashi T, Imai M
Japan Synthetic Rubber Co., Ltd. Tsukuba Research Laboratory.
Kansenshogaku Zasshi. 1995 Sep;69(9):957-62. doi: 10.11150/kansenshogakuzasshi1970.69.957.
PCR mediated detection of HIV-DNA has been widely used. However, compared with traditional immunological diagnoses, the extraction of DNA is a laborious and time consuming step. We have developed a procedure for the efficient isolation and concentration of HIV-DNA from cell lysates. We report here a novel method by one can recover HIV-DNA in a small volume (approximately 50 microliters) of solution from a large volume of crude cell lysate which contains as few as several copies. The method uses the specific hybridization of HIV-DNA to HIV probe-DNA particles. This prior enrichment augmented the sensitivity in the detection of HIV-DNA by PCR, and allows us to make a diagnosis even if the specimen contained an extremely low copy number of HIV-DNA molecules in a large volume, which would have otherwise resulted in false-negative data with the conventional extraction method. The method also enables the examination of 100 individual blood specimens in a combined form. Thus, the application of the present enrichment procedure with HIV probe-DNA particles should reduce the labor and cost of HIV diagnosis, since the HIV positive samples represent a very minor group of people among specimens subjected to clinical laboratory tests, and particularly, among blood samples voluntarily donated to be used for transfusions.
聚合酶链反应（PCR）介导的HIV-DNA检测已被广泛应用。然而，与传统的免疫诊断方法相比，DNA提取是一个费力且耗时的步骤。我们开发了一种从细胞裂解物中高效分离和浓缩HIV-DNA的方法。在此，我们报告一种新方法，通过该方法可以从大量粗细胞裂解物中，在小体积（约50微升）溶液中回收HIV-DNA，即使粗细胞裂解物中仅含有几个拷贝数。该方法利用HIV-DNA与HIV探针-DNA颗粒的特异性杂交。这种预先富集提高了PCR检测HIV-DNA的灵敏度，即使样本在大量溶液中含有极低拷贝数的HIV-DNA分子，使用传统提取方法可能会产生假阴性数据，但该方法仍能使我们做出诊断。该方法还能够以组合形式检测100份个体血液样本。因此，应用目前这种含HIV探针-DNA颗粒的富集方法应可降低HIV诊断的劳动强度和成本，因为在接受临床实验室检测的样本中，尤其是在自愿捐献用于输血的血液样本中，HIV阳性样本只占极少数人群。
