多标记免疫荧光染色及多光谱成像技术在组织学研究中的应用

摘要

免疫组织化学染色及成像分析是研究组织形态和组织原位抗原表达不可或缺的检测技术,广泛应用于临床病理诊断和医学及生物学研究的各个领域.组织切片样本中蕴含着丰富的信息,但是受制于传统单标记免疫组织化学染色方法的限制,通常只能对组织中的一到两种抗原进行染色分析,而且定量结果的判读往往依赖于肉眼观测,缺乏客观标准.随着蛋白组学的发展,对现代组织学分析提出了更高的要求,例如不同的蛋白共表达和共定位分析、低丰度分子的检测、异质性分析、细胞表型统计乃至复杂组织微环境的描绘等,都需要在同一张组织切片样本上同时检测多种靶标分子.这对于理解组织微环境中各种细胞间的关系,推演信号通路上下游蛋白表达的关系,制定临床诊断和治疗方案都有着非常重要的意义.本文介绍一种基于酪胺信号放大(TSA)技术衍生而来的多标记免疫荧光染色方法,能够在同一组织切片样本上复染7种以上抗原并进行区别标记,配合光谱成像技术和定量分析软件,能够将组织中蕴含的丰富信息准确的呈现出来.这套流程化的分析方案为临床诊断和基础科研提供了更高精度和更可靠的组织学数据,将免疫组织化学分析的技术水平提升到一个新的高度.%Immunohistochemical staining and imaging analysis technology is indispensable for the study of tissue morphology and in situ antigen expression.It is widely used in various fields of clinical pathology diagnosis and biological research.The tissue slice sample contains a lot of information,but due to the restriction of traditional single-labeled immunohistochemical staining method,only one or two target antigens can be detected at same time.The quantitative interpretation of the results is often dependent on naked eye observation and lacks objective criteria.With the development of proteomics,there are increasing demands on more sophisticated modem histological analysis,such as different protein co-expression and co-localization analysis,low abundance molecular detection,heterogeneity analysis,cell phenotype statistics,and complex tissue microenvironment description.These features are essential in detecting a variety of target molecules in the same tissue slice simultaneously so that researchers can understand the relationship among various cells in tissue microenvironment,predict the relationship between upstream and downstream protein expression,and develop clinical diagnosis and treatment regimen.This review presents a multi-labeled immunofluorescence staining method based on tyramine signal amplification (TSA) technology.It allows more than seven different markers on the same tissue slice using different colored dyes combined with spectral imaging and quantitative analysis software,therefore the abundant in situ tissue information can be accurately presented.This set of process-oriented analysis can provide more accurate and reliable histological data for clinical diagnosis and scientific research,and bring the immunohistochemical analysis to a higher level.