An Emerging Approach for Parallel Quantification of Intracellular Protozoan Parasites and Host Cell Characterization Using TissueFAXS Cytometry
An Emerging Approach for Parallel Quantification of Intracellular Protozoan Parasites and Host Cell Characterization Using TissueFAXS Cytometry
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DOI：
10.1371/journal.pone.0139866
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发表时间：
2015-10-21
期刊：
影响因子：
3.7
通讯作者：
Ritter, Uwe
中科院分区：
文献类型：
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作者：
Schmid, Maximilian;Dufner, Bianca;Ritter, Uwe
宿主-病原体相互作用的表征是微生物学和免疫学学科的基本方法。人们普遍认为，宿主细胞在吞噬病原体后开始改变其表型。实时PCR或ELISA等技术被用于表征编码与病原体消除或免疫逃逸机制相关的蛋白质的基因。大多数此类研究都是在体外使用原代宿主细胞或细胞系进行的。因此，用这种方法产生的数据反映了从培养的所有细胞中恢复的全局RNA表达或蛋白质量。在实验条件下，当所有宿主细胞都含有等量的病原体时，这是合理的。然而，吞噬细胞对病原体的摄取是不同步的。因此，宿主细胞含有不同数量的病原体，可能导致不同的病原体诱导的蛋白质生物合成。因此，我们建立了一种能够使用基于免疫荧光的高通量分析来检测和量化相应宿主细胞中病原体数量的技术。与感兴趣的分子的多色染色配对，现在可以分析宿主细胞群的感染概况和宿主细胞因寄生虫负荷而产生的相应表型。
Characterization of host-pathogen interactions is a fundamental approach in microbiological and immunological oriented disciplines. It is commonly accepted that host cells start to change their phenotype after engulfing pathogens. Techniques such as real time PCR or ELISA were used to characterize the genes encoding proteins that are associated either with pathogen elimination or immune escape mechanisms. Most of such studies were performed in vitro using primary host cells or cell lines. Consequently, the data generated with such approaches reflect the global RNA expression or protein amount recovered from all cells in culture. This is justified when all host cells harbor an equal amount of pathogens under experimental conditions. However, the uptake of pathogens by phagocytic cells is not synchronized. Consequently, there are host cells incorporating different amounts of pathogens that might result in distinct pathogen-induced protein biosynthesis. Therefore, we established a technique able to detect and quantify the number of pathogens in the corresponding host cells using immunofluorescence-based high throughput analysis. Paired with multicolor staining of molecules of interest it is now possible to analyze the infection profile of host cell populations and the corresponding phenotype of the host cells as a result of parasite load.