Levitz S M, Diamond R D
J Infect Dis. 1985 Nov;152(5):938-45. doi: 10.1093/infdis/152.5.938.
A rapid colorimetric assay for viability of fungi that uses the tetrazolium salt MTT is described. All live species of a wide range of medically important fungi that were tested reduced MTT, with resultant intracellular formation within several hours of clearly visible purple crystals of MTT-formazan. The assay proved particularly useful in measuring viability of individual hyphae; greater than 99% of live Aspergillus fumigatus and Rhizopus oryzae hyphae stained purple when incubated with MTT compared with 0% of hyphae killed by any of a variety of methods. The MTT method for measuring fungal viability closely correlated with a slide-culture technique when killing of A. fumigatus hyphae by graded concentrations of hydrogen peroxide was compared. MTT-formazan was easily extracted from the fungi by alcohol, a procedure allowing spectrophotometric quantification of fungal killing in both cell-free and cellular systems. A linear relation between inocula and MTT reduction of several fungi, including Candida albicans blastospores and A. fumigatus hyphae, was demonstrated. Human leukocytes killed 80.3% of C. albicans blastospores as measured by the MTT assay, compared with 70.5% as measured by dilutions and pour plates. Killing of A. fumigatus hyphae by human leukocytes was also quantitated by using the MTT assay.
本文描述了一种使用四氮唑盐MTT的快速比色法来检测真菌的活力。所测试的多种具有医学重要性的真菌的所有活菌种均能使MTT还原，在数小时内细胞内形成清晰可见的紫色MTT-甲臜晶体。该检测方法在测量单个菌丝的活力方面特别有用；与通过多种方法杀死的0%的菌丝相比，当与MTT一起孵育时，超过99%的活烟曲霉和米根霉菌丝被染成紫色。当比较不同浓度的过氧化氢对烟曲霉菌丝的杀伤作用时，MTT法测量真菌活力与玻片培养技术密切相关。MTT-甲臜很容易用酒精从真菌中提取出来，这一过程允许在无细胞和细胞系统中对真菌杀伤进行分光光度法定量。证明了包括白色念珠菌芽生孢子和烟曲霉菌丝在内的几种真菌的接种量与MTT还原之间存在线性关系。通过MTT检测法测量，人类白细胞杀死了80.3%的白色念珠菌芽生孢子，而通过稀释和平板倾注法测量的结果为70.5%。还使用MTT检测法定量了人类白细胞对烟曲霉菌丝的杀伤作用。
