Construction of a multiple fluorescence labelling system for use in co-invasion studies of Listeria monocytogenes.
Construction of a multiple fluorescence labelling system for use in co-invasion studies of Listeria monocytogenes.
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DOI：
10.1186/1471-2180-6-86
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发表时间：
2006-10-03
期刊：
影响因子：
4.2
通讯作者：
Licht TR
中科院分区：
文献类型：
--
作者：
Andersen JB;Roldgaard BB;Lindner AB;Christensen BB;Licht TR
现有的毒力模型往往难以应用于单增李斯特菌入侵潜力的定量比较。基于细胞系的体外测定之间的孔间差异实际上是不可避免的，并且当使用动物模型时，个体动物之间的差异是观察到的感染能力的大偏差的原因。解决这一问题的一种方法是进行毒力研究，作为2种或2种以上菌株之间的竞争试验。然而，这需要能够容易区分不同菌株的侵袭中性标记。荧光标记系统，允许可视化和识别单个L。单核细胞增多症细胞以及集落以非破坏性的方式，开发。五种不同的荧光标记是可用的，并允许通过使用荧光显微镜在单细胞水平上同时视觉区分三种不同标记的菌株。90%以上的L.单核细胞增多症宿主细胞保持荧光标记40代。荧光标记并没有改变L.在传统的Caco-2细胞侵袭测定中，通过对单核细胞增多性细胞的荧光标记，可以在细胞内携带不同荧光标记的侵袭细菌之间进行视觉区分。所构建的荧光标记体系稳定、使用方便、不影响L.在Caco-2细胞测定中，单核细胞增多症的细胞因子可以被检测到，并且允许在这些细胞中内化后区分不同标记的细菌。
Existing virulence models are often difficult to apply for quantitative comparison of invasion potentials of Listeria monocytogenes. Well-to-well variation between cell-line based in vitro assays is practically unavoidable, and variation between individual animals is the cause of large deviations in the observed capacity for infection when animal models are used. One way to circumvent this problem is to carry out virulence studies as competition assays between 2 or more strains. This, however, requires invasion-neutral markers that enable easy discrimination between the different strains. A fluorescent marker system, allowing visualization and identification of single L. monocytogenes cells as well as colonies in a non-destructive manner, was developed. Five different fluorescent labels are available, and allowed simultaneous visual discrimination between three differently labelled strains at the single cell level by use of fluorescence microscopy. More than 90% of the L. monocytogenes host cells maintained the fluorescence tags for 40 generations. The fluorescence tags did not alter the invasive capacity of the L. monocytogenes cells in a traditional Caco-2 cell invasion assay, and visual discrimination between invaded bacteria carrying different fluorescent labels inside the cells was possible. The constructed fluorescent marker system is stable, easy to use, does not affect the virulence of L. monocytogenes in Caco-2 cell assays, and allows discrimination between differently labelled bacteria after internalization in these cells.
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影响因子：
6.7
作者：
Pentecost, Mickey;Otto, Glen;Theriot, Julie A;Amieva, Manuel R
通讯作者：
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DOI：
10.1073/pnas.97.22.11984
发表时间：
2000-10-24
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作者：
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