利用生物发光分析技术快速检测微生物的方法学研究

STUDY ON THE RAPID BIOLUMINESCENCE ASSAY METHOD FOR THE DETECTION OF BACTERIA

阻碍三磷酸腺苷（ＡＴＰ）－虫萤光素酶发光体系用于微生物快速检测的主要因素之一，在于使用了不适当的非微生物ＡＴＰ清除方法和微生物ＡＴＰ提取方法．文章对非微生物ＡＴＰ的清除和微生物ＡＴＰ提取方法进行了实验研究．选择ＴｒｉｔｏｎＸ－１００作为非微生物ＡＴＰ淬取剂．Ａｐｙｒａｓｅ作为非微生物ＡＴＰ清除剂．三氯醋酸作为微生物ＡＴＰ的淬取剂．观察了淬取剂及清除剂浓度对淬取效果及ＡＴＰ分析的影响．最佳工作浓度分别为：ＴｒｉｔｏｎＸ－１０００．１５％、ａｐｙｒａｓｅ０．１％、三氯醋酸１．５％．对超声、加热超声、加热、三氯醋酸、氯仿淬取微生物ＡＴＰ的方法作了对比研究，认为三氯醋酸方法更简便。

One of major factors that have hampered bioluminescent ATP techniques application to biomass assays is the improper removal methods of somatic ATP and inadequate microbial ATP extraction methods that have been used in the ATP rapid microbiology. The experimental study of non microbial ATP removal methods and the microbial ATP extraction methods has been done. Triton X 100 and apyrase are used as extractant and removal reagent of somatic ATP. Trichlorracetic acid is used to extract microbial ATP. The optimum concentration of them has successively been determined as 0.15%, 0.10% and 1.50%. The comparative research of microbial ATP extraction methods such as ultrasonic disintegration, boiling, ultrasonic boiling, TCA and chloroform has been done. The conclusion is that the trichloroacetic acid is preferable to other extraction methods. The method is simple and easy to be widely used.