MALDI-TOF-MS用于微生物鉴定的影响因素分析
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摘要
影响MALDI-TOF-MS微生物鉴定的因素有很多。本研究主要从微生物培养基、培养状态、菌体前处理、上样量等方面进行MALDI-TOF-MS操作技术和方法的探讨。研究结果显示:进行MALDI-TOF-MS细菌鉴定,培养时间在4～24 h之间为佳;尽量选择无色、不含盐的液体培养基;0.1μL菌液是最低鉴定菌量;上样量对结果无影响;对于菌体前处理,除按照标准化前处理程序操作外,也可将菌体加dd H2O制成混悬液进行点靶鉴定,这样更加简便。本研究有助于减少MALDI-TOF-MS操作盲目性,提高鉴定质量,利于操作的规范化和简便化。
Various factors could influence the effect of MALDI-TOF-MS in microorganism identification. In this study,the approaches and techniques of MALDI-TOF-MS from aspects of microbiological medium,bacteria incubation state,bacteria pre-treatment and sample loading quantity were summarized and discussed. According to the research results,the period of bacterium incubation of 4 to 24 hours was the best;colorless and salt-free medium was preferable;the lowest ascertainable bacteria solution was 0.1 μL;the sample loading quantity had no effect on identification result. Besides,for the standard pre-treatment procedures,a handier way was to add ddH2 O into the bacteria to make suspension and conduct target identification. This study would contribute to reduce the operational blindness of MALDI-TOF-MS,enhance the identification quality and facilitate the standardization and simplification of operation.
Various factors could influence the effect of MALDI-TOF-MS in microorganism identification. In this study,the approaches and techniques of MALDI-TOF-MS from aspects of microbiological medium,bacteria incubation state,bacteria pre-treatment and sample loading quantity were summarized and discussed. According to the research results,the period of bacterium incubation of 4 to 24 hours was the best;colorless and salt-free medium was preferable;the lowest ascertainable bacteria solution was 0.1 μL;the sample loading quantity had no effect on identification result. Besides,for the standard pre-treatment procedures,a handier way was to add ddH2 O into the bacteria to make suspension and conduct target identification. This study would contribute to reduce the operational blindness of MALDI-TOF-MS,enhance the identification quality and facilitate the standardization and simplification of operation.
引文
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[7]陈爱平,陈建辉,杨劲松,等.分子生物学技术在肠致泻性大肠杆菌诊断中的应用[J].中国人兽共患病学报,2011,27(9):808-811.
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[9]白瑶,叶淑瑶,闫韶飞,等.2015年我国食源性副溶血性弧菌毒力基因及耐药特征研究[J].食品安全质量检测学报,2017,9(4):884-889.
[2]孙宗科,张伟,陈西平.应用飞行时间质谱仪快速鉴定细菌的初步研究[J].卫生研究,2004,33(5):552-554.
[3]HOLLAND R D,DUFFY C R,RAFII R,et a1.Identification of bacterial proteins observed in MALDITOF mass spectra from whole cells[J].Analytical chemistry,1999,71(15):3226-3230.
[4]ARNOLD R J,KARTY J A,ELLINGTON A D,et a1.Monitoring the growth of a bacteria culture by MALDI-MS of whole cells[J].Analytical chemistry,1999,71(10):1990-1996.
[5]VELUSAMY V,ARSHAK K,KOROSTYNSKA O,et al.An overview of foodborne pathogen detection:in the perspective of biosensors[J].Biotechnology advances,2010,28(2):232-254.
[6]徐淑菲,孔繁德.MALDI Biotyper高通量微生物鉴定系统在进口甲鱼蛋检疫中的初步应用[J].厦门检验检疫科技,2010(2):30-35.
[7]陈爱平,陈建辉,杨劲松,等.分子生物学技术在肠致泻性大肠杆菌诊断中的应用[J].中国人兽共患病学报,2011,27(9):808-811.
[8]黄文宇,柳陈坚.食源性沙门氏菌检测方法的研究进展[J].生物技术,2009,19(3):95-98.
[9]白瑶,叶淑瑶,闫韶飞,等.2015年我国食源性副溶血性弧菌毒力基因及耐药特征研究[J].食品安全质量检测学报,2017,9(4):884-889.