Using Imaging Flow Cytometry to Quantify Neutrophil Phagocytosis.
Using Imaging Flow Cytometry to Quantify Neutrophil Phagocytosis.
复制标题
使用成像流式细胞术定量中性粒细胞吞噬作用。
DOI：
10.1007/978-1-0716-0154-9_10
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
Criss,AlisonK
中科院分区：
文献类型：
--
作者：
Smirnov,Asya;Solga,MichaelD;Lannigan,Joanne;Criss,AlisonK
Neutrophils are professional phagocytes that are important for innate host defenses against pathogens and resolution of inflammation. Traditionally, the phagocytic capacity of neutrophils was quantified by enumeration of cells containing either internalized or bound bacteria or other cargo from a series of microscopic images. Here we describe an imaging flow cytometry-based protocol and analysis method for quantifying the binding and uptake ofNeisseria gonorrhoeaeby primary adherent human neutrophils. Imaging flow cytometry combines the capacity for quantitative, high-throughput analysis of tens of thousands of cells per condition, with the imaging power of fluorescence microscopy. Here, all bacteria are labeled with Tag-it Violet™ and bound bacteria are differentially stained with a DyLight™ 650-conjugated antibody. Images are analyzed using spot count and other algorithms. Outputs include the percent of neutrophils associated with bacteria, the percent of neutrophils with internalized bacteria, and the percent of internalized bacteria. This basic protocol can be adapted to a variety of particle types and can be used for multiplex analysis in combination with staining for different neutrophil surface and intracellular markers.
