生物荧光法检测结核分枝杆菌ATP含量  

Bioluminescence assay of Mycobacterium tuberculosis

作　　者：张虹妍[1] 石瑞如[1] 史慧敏[1] 孙芳[1] 秦殊[1] 张霞[1] 张国龙[1] 

机构地区：[1]河南省胸科医院,郑州450008

出　　处：《医药论坛杂志》2011年第17期1-2,5,共3页Journal of Medical Forum

基　　金：河南省医学科技攻关项目(编号2006149)

摘　　要：目的建立生物荧光技术检测结核分枝杆菌ATP含量的方法并观察异烟肼的抗结核作用。方法结核分枝杆菌标准株H37Rv的菌悬液,高温裂解,用生物荧光技术检测三磷酸腺苷(ATP)含量。条件稳定后,观察菌数和ATP浓度的关系,以及加入不同浓度的异烟肼后1周之内ATP的动态变化。结果菌液加入裂解液后100℃煮沸8min,迅速降至室温,加入荧光素酶,3～5min时测荧光值。随异烟肼浓度增加,ATP量下降,培养至第7d时,异烟肼的敏感性可以初步进行判断。结论所建立的生物荧光法检测结核分枝杆菌ATP含量,具有简便、快速、敏感、可重复性强等特点,有望成为结核菌体外药敏试验的一种有用的方法。Objective To monitor mycobacterial growth by bioluminescence assay of mycobacterial ATP and to analyze the susceptibility of isoniazid.Methods ATP was extracted from the bacterial suspension prepared from Mycobacterium tuberculosis H37Rv and measured by bioluminescent assay.The ATP contents of H37Rv inoculated into the Middlebrook 7H9 broth medium containing antituberculosis agents were measured at days of 0,3,5 and 7.Results The highest relative light units（RLU） was obtained when ATP was extracted with lysis buffer at 100℃ boiling for 8 minutes and then decreased quickly to room temperature.3 to 5 minutes after adding biluminescence enzyme was found to be the proper time for RLU test.The control culture showed the time-dependent increase in the RLU values,while cultures supplemented with antimicrobial agents reduced their ATP contents concomitant with the concentrations of drugs.Conclusions The bioluminescence assay of mycobacterial ATP established in this work is simple,rapid,sensitive and highly reproducible.

关 键 词：结核菌 三磷酸腺苷 生物发光法 

分 类 号：R927.1[医药卫生—药学]