Validation of CyTOF Against Flow Cytometry for Immunological Studies and Monitoring of Human Cancer Clinical Trials
Validation of CyTOF Against Flow Cytometry for Immunological Studies and Monitoring of Human Cancer Clinical Trials
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DOI：
10.3389/fonc.2019.00415
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发表时间：
2019-05-17
影响因子：
4.7
通讯作者：
Brooks, David G.
中科院分区：
文献类型：
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作者：
Gadalla, Ramy;Noamani, Babak;Brooks, David G.
流式细胞术是一种广泛应用于癌症和其他疾病的探索性免疫分析和生物标志物发现的方法。然而，流式细胞术受到可以同时分析的参数数量的限制，严重限制了其实用性。最近，质谱细胞术（CyTOF）的出现实现了对免疫系统的多维和公正的检查，允许同时询问大量参数。这对于深入询问免疫反应非常重要，特别是当样本量有限时（例如肿瘤）。我们的目标是比较CyTOF与流式细胞术的准确性和可重复性，作为癌症临床试验中人类PBMC和肿瘤组织的可靠分析工具。我们开发了40+参数的CyTOF面板，并证明与流式细胞术相比，CyTOF结合细胞分化、功能、活化和耗竭的标志物产生类似的细胞谱系定量，用于新鲜和有活力的冷冻PBMC或肿瘤组织。此外，我们提供了一个协议，使可靠的定量CyTOF低到低数量的输入人类细胞，一种方法，是特别重要的细胞数量是有限的。因此，我们验证了CyTOF作为一种准确的方法，在人类肿瘤组织中进行高维分析，并利用低细胞数进行后续的免疫学研究和癌症临床试验。
Flow cytometry is a widely applied approach for exploratory immune profiling and biomarker discovery in cancer and other diseases. However, flow cytometry is limited by the number of parameters that can be simultaneously analyzed, severely restricting its utility. Recently, the advent of mass cytometry (CyTOF) has enabled high dimensional and unbiased examination of the immune system, allowing simultaneous interrogation of a large number of parameters. This is important for deep interrogation of immune responses and particularly when sample sizes are limited (such as in tumors). Our goal was to compare the accuracy and reproducibility of CyTOF against flow cytometry as a reliable analytic tool for human PBMC and tumor tissues for cancer clinical trials. We developed a 40+ parameter CyTOF panel and demonstrate that compared to flow cytometry, CyTOF yields analogous quantification of cell lineages in conjunction with markers of cell differentiation, function, activation, and exhaustion for use with fresh and viably frozen PBMC or tumor tissues. Further, we provide a protocol that enables reliable quantification by CyTOF down to low numbers of input human cells, an approach that is particularly important when cell numbers are limiting. Thus, we validate CyTOF as an accurate approach to perform high dimensional analysis in human tumor tissue and to utilize low cell numbers for subsequent immunologic studies and cancer clinical trials.