A method for high throughput determination of viable bacteria cell counts in 96-well plates.
A method for high throughput determination of viable bacteria cell counts in 96-well plates.
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高吞吐量测定96孔板中可行细菌计数的一种方法。
DOI：
10.1186/1471-2180-12-259
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发表时间：
2012-11-13
期刊：
影响因子：
4.2
通讯作者：
Rahme LG
中科院分区：
文献类型：
--
作者：
Hazan R;Que YA;Maura D;Rahme LG
有几种方法用于定量细菌细胞，每种方法都有优点和缺点。最常见的方法是细菌平板法，其优点是通过菌落形成单位（CFU）计数进行活细胞评估，但不太适合高通量筛选（HTS）。另一方面，分光光度法适用于HTS应用，但不能区分死细菌和活细菌，灵敏度低。在这里，我们报告了一种细菌细胞计数方法，称为开始生长时间（SGT），允许快速和连续定量的绝对或相对数量的活细胞在细菌培养物中的高通量的方式。我们将定量聚合酶链反应（qPCR）计算的方法与先前描述的细菌生长测定的定性方法相结合，以开发一种改进的定量方法。我们表明，SGT仅检测活细菌，并且灵敏度足以区分40和400个细胞/mL。SGT基于生长细胞培养物达到阈值所需的再生长时间，以及该时间与初始接种物中的细胞数量成比例的概念。我们展示了SGT的几个应用，包括评估抗生素对细胞活力的影响和确定细胞群中抗生素耐受亚群的比例。SGT结果与CFU计数结果无显著差异。SGT是一种相对快速、高度灵敏、可重复且不费力的方法，可用于HTS环境中，以纵向评估细菌细胞培养物中的活细胞。
There are several methods for quantitating bacterial cells, each with advantages and disadvantages. The most common method is bacterial plating, which has the advantage of allowing live cell assessment through colony forming unit (CFU) counts but is not well suited for high throughput screening (HTS). On the other hand, spectrophotometry is adaptable to HTS applications but does not differentiate between dead and living bacteria and has low sensitivity. Here, we report a bacterial cell counting method termed Start Growth Time (SGT) that allows rapid and serial quantification of the absolute or relative number of live cells in a bacterial culture in a high throughput manner. We combined the methodology of quantitative polymerase chain reaction (qPCR) calculations with a previously described qualitative method of bacterial growth determination to develop an improved quantitative method. We show that SGT detects only live bacteria and is sensitive enough to differentiate between 40 and 400 cells/mL. SGT is based on the re-growth time required by a growing cell culture to reach a threshold, and the notion that this time is proportional to the number of cells in the initial inoculum. We show several applications of SGT, including assessment of antibiotic effects on cell viability and determination of an antibiotic tolerant subpopulation fraction within a cell population. SGT results do not differ significantly from results obtained by CFU counts. SGT is a relatively quick, highly sensitive, reproducible and non-laborious method that can be used in HTS settings to longitudinally assess live cells in bacterial cell cultures.
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影响因子：
6.7
作者：
Hazan R;He J;Xiao G;Dekimpe V;Apidianakis Y;Lesic B;Astrakas C;Déziel E;Lépine F;Rahme LG
通讯作者：
Rahme LG
影响因子：
7
作者：
Heid, CA;Stevens, J;Williams, PM
通讯作者：
Williams, PM
影响因子：
6.7
作者：
Kesarwani, Meenu;Hazan, Ronen;Rahme, Laurence G.
通讯作者：
Rahme, Laurence G.
DOI：
10.1073/pnas.251465298
发表时间：
2001-12-04
影响因子：
11.1
作者：
Cao, H;Krishnan, G;Rahme, LG
通讯作者：
Rahme, LG
DOI：
10.1073/pnas.0307694100
发表时间：
2004-02-03
影响因子：
11.1
作者：
Déziel, E;Lépine, F;Rahme, LG
通讯作者：
Rahme, LG