Flow Cytometry as an Improved Method for the Titration of Chlamydiaceae and Other Intracellular Bacteria
Flow Cytometry as an Improved Method for the Titration of Chlamydiaceae and Other Intracellular Bacteria
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DOI：
10.1002/cyto.a.22822
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发表时间：
2016-05-01
期刊：
影响因子：
3.7
通讯作者：
Meurens, F.
中科院分区：
文献类型：
--
作者：
Kaser, T.;Pasternak, J. A.;Meurens, F.
衣原体科是细胞内细菌的一个家族，引起一系列不同的病理结果。最具破坏性的人类疾病是眼部感染C。沙眼导致失明，生殖器感染导致盆腔炎，并有长期后遗症，包括不孕症和慢性盆腔疼痛。为了能够比较研究宿主衣原体相互作用的实验室之间的实验，必须测定感染滴度。衣原体的滴度测定最常通过用衣原体的连续稀释液感染的宿主细胞的显微镜检查来进行。然而，其他方法，包括荧光ELISpot（Fluorospot）和DNA芯片扫描技术也被提出来计数衣原体感染的细胞。对于病毒，流式细胞术被认为是标准滴定方法的上级替代方法。在这项研究中，我们比较了使用流式细胞术与显微镜和荧光点滴定C。suis作为其它胞内细菌的代表。通过Fluorospot进行滴度测定是不可靠的，而通过显微镜进行滴定导致16264稀释度的线性读出范围和中等重现性，研究者内和研究者之间的标准偏差可接受。相比之下，流式细胞术具有1，024个稀释度的广泛线性读出范围，并且在这些方法的基础训练下具有最低的标准偏差。此外，与显微镜相比，流式细胞术更快，材料成本更低。流式细胞术为细胞内细菌（如C。猪。(C)2016年国际细胞计数促进学会
Chlamydiaceae is a family of intracellular bacteria causing a range of diverse pathological outcomes. The most devastating human diseases are ocular infections with C. trachomatis leading to blindness and genital infections causing pelvic inflammatory disease with long-term sequelae including infertility and chronic pelvic pain. In order to enable the comparison of experiments between laboratories investigating hostchlamydia interactions, the infectious titer has to be determined. Titer determination of chlamydia is most commonly performed via microscopy of host cells infected with a serial dilution of chlamydia. However, other methods including fluorescent ELISpot (Fluorospot) and DNA Chip Scanning Technology have also been proposed to enumerate chlamydia-infected cells. For viruses, flow cytometry has been suggested as a superior alternative to standard titration methods. In this study we compared the use of flow cytometry with microscopy and Fluorospot for the titration of C. suis as a representative of other intracellular bacteria. Titer determination via Fluorospot was unreliable, while titration via microscopy led to a linear read-out range of 16264 dilutions and moderate reproducibility with acceptable standard deviations within and between investigators. In contrast, flow cytometry had a vast linear read-out range of 1,024 dilutions and the lowest standard deviations given a basic training in these methods. In addition, flow cytometry was faster and material costs were lower compared to microscopy. Flow cytometry offers a fast, cheap, precise, and reproducible alternative for the titration of intracellular bacteria like C. suis. (C) 2016 International Society for Advancement of Cytometry