Parra Edwin Roger, Jiang Mei, Solis Luisa, Mino Barbara, Laberiano Caddie, Hernandez Sharia, Gite Swati, Verma Anuj, Tetzlaff Michael, Haymaker Cara, Tamegnon Auriole, Rodriguez-Canales Jaime, Hoyd Clifford, Bernachez Chantale, Wistuba Ignacio
Department of Translational Molecular Pathology, the University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
Department of Pathology, Division of Pathology and Laboratory Medicine. The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA.
Cancers (Basel). 2020 Jan 21;12(2):255. doi: 10.3390/cancers12020255.
In the development of a multiplex immunofluorescence (IF) platform and the optimization and validation of new multiplex IF panels using a tyramide signal amplification system, several technical requirements are important for high-quality staining, analysis, and results. The aim of this review is to discuss the basic requirements for performing multiplex IF tyramide signal amplification (TSA) in formalin-fixed, paraffin-embedded cancer tissues to support translational oncology research. Our laboratory has stained approximately 4000 formalin-fixed, paraffin-embedded tumor samples using the multiplex IF TSA system for immune profiling of several labeled biomarkers in a single slide to elucidate cancer biology at a protein level and identify therapeutic targets and biomarkers. By analyzing several proteins in thousands of cells on a single slide, this technique provides a systems-level view of various processes in various tumor tissues. Although this technology shows high flexibility in cancer studies, it presents several challenges when applied to study different histology cancers. Our experience shows that adequate antibody validation, staining optimization, analysis strategies, and data generation are important steps for generating quality results. Tissue management, fixation procedures, storage, and cutting can also affect the results of the assay and must be standardized. Overall, this method is reliable for supporting translational research given a precise, step-by-step approach.
在开发多重免疫荧光（IF）平台以及使用酪胺信号放大系统优化和验证新的多重IF检测板时，对于高质量的染色、分析和结果而言，有几个技术要求至关重要。本综述的目的是讨论在福尔马林固定、石蜡包埋的癌症组织中进行多重IF酪胺信号放大（TSA）的基本要求，以支持转化肿瘤学研究。我们实验室已使用多重IF TSA系统对约4000个福尔马林固定、石蜡包埋的肿瘤样本进行染色，以便在一张载玻片上对几种标记生物标志物进行免疫分析，从而在蛋白质水平阐明癌症生物学并识别治疗靶点和生物标志物。通过在一张载玻片上分析数千个细胞中的几种蛋白质，该技术提供了各种肿瘤组织中各种过程的系统级视图。尽管这项技术在癌症研究中显示出高度的灵活性，但在应用于研究不同组织学类型的癌症时仍存在一些挑战。我们的经验表明，充分的抗体验证、染色优化、分析策略和数据生成是获得高质量结果的重要步骤。组织管理、固定程序、储存和切片也会影响检测结果，必须进行标准化。总体而言，鉴于采用精确的分步方法，该方法对于支持转化研究是可靠的。
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