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小木虫论坛-学术科研互动平台 » 生物医药区 » 生物科学 » 求助 细胞流式检测循环肿瘤细胞方法
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sun332008

金虫 (小有名气)

[求助] 求助 细胞流式检测循环肿瘤细胞方法 已有1人参与

求助   我想请教 细胞流式检测循环肿瘤细胞具体实验方法步骤，提供一下相关这方面的文献也好，谢谢。
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2021， 向钱看， 向厚赚， 做一个快乐的小富婆。 加油呀！
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wotaoyan

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从文献中复制的，希望对你有帮助，参考文献：Cetuximab combined with natural killer cells therapy: an alternative to chemoradiotherapy for patients with advanced non-small cell lung cancer (NSCLC)，Am J Cancer Res 2018;8(5):879-891
Peripheral blood (7 mL) was obtained from patients on day 1 and day 90 for the detection of CTC levels. MNCs from peripheral blood  were separated using LymphoprepTM (Haoyang Biologicals) density gradient centrifugation from buffy coats and were washed twice with sterile Hank’s balanced salt solution (Life Technologies, Carlsbad, CA, USA). Isolated cells were enriched via binding to magnetic CD326 (EpCAM) MicroBeads (Miltenyi Biotech, Bergisch Gladbach, Germany) using magnetic activated cell sorting. Then, enriched isolated cells were labelled with monoclonal antibodies targeting the epithelial cell antigens CD45, CD326 as well as cytokeratins 8, 18 and 19 (Miltenyi Biotech) and incubated in the dark for 12 min at room temperature. The monoclonal antibodies (10 μL) anti-CD45-PE, Ep-CAM-APC,and cytokeratins 8, 18, and 19-FITC were added per 7.5 mL of whole blood. Cell pellets were resuspended in  500  μL  PBS  and  enumerated  by  a FACSCantoTM II (Becton Dickinson, Franklin  Lakes,  NJ,  USA)  flow  cytometer with a CD45-/CK+/CD326+ gating strategy. The absolute number of CD45-, CK+, and CD326+ cells on FACSCantoTM II was used to measure CTC levels 。
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