CTC identification and characterization in breast cancer using K/V protocol and imaging flow cytometry - part 4

This project is a part of bigger study aiming to identify and characterize circulating tumor cells (CTCs) in peripheral blood of cancer patients. Here, specificaly it focused on CTCs analysis in 54 breast cancer patients. Fraction of peripheral blood mononuclear cells putatively containing CTCs was isolated using density gradient centrifugation. Pan-keratin/vimentin/CD45/CD31/DAPI immunofluorescent staining was performer and DAPI+ or DAPI+CD45/CD31- objects were collected using Amnis® ImageStream®XMk II, Cytek® Biosciences, Inc (lasers: 405 nm, 488 nm, 642 nm, magnification 40x, slow speed flow).本项目隶属于一项旨在鉴定并表征癌症患者外周血中循环肿瘤细胞（circulating tumor cells, CTCs）的大型研究。本研究具体针对54名乳腺癌患者的循环肿瘤细胞展开分析。研究人员采用密度梯度离心法，分离出疑似含有循环肿瘤细胞的外周血单个核细胞组分。随后采用广谱角蛋白/波形蛋白/CD45/CD31/DAPI进行免疫荧光染色，并使用赛泰克生物科学公司（Cytek® Biosciences, Inc）的Amnis® ImageStream®XMk II成像流式细胞仪采集目标细胞：即DAPI阳性或DAPI阳性且CD45/CD31阴性的细胞，实验配置参数为激光波长405 nm、488 nm、642 nm，放大倍数40×，采用低速流式模式。