A recommended and verified procedure for in situ tryptic digestion of formalin-fixed paraffin-embedded tissues for analysis by matrix-assisted laser desorption/ionization imaging mass spectrometry
A recommended and verified procedure for in situ tryptic digestion of formalin-fixed paraffin-embedded tissues for analysis by matrix-assisted laser desorption/ionization imaging mass spectrometry
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DOI：
10.1002/jms.4384
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发表时间：
2019-08-01
影响因子：
2.3
通讯作者：
Caprioli, Richard M.
中科院分区：
文献类型：
--
作者：
Judd, Audra M.;Gutierrez, Danielle B.;Caprioli, Richard M.
基质辅助激光解吸/电离成像质谱（MALDI IMS）是一种独特的分子成像技术，能够在保留其原位位置的空间信息的同时，对蛋白质、脂质和代谢物进行非靶向测量。这种强大的能力组合有可能为分子组织学领域带来丰富的知识。将这种创新的研究工具转化为临床实验室，需要开发可靠的样品制备方案，用于分析福尔马林固定石蜡包埋（FFPE）组织中的蛋白质，这是临床病理学中的标准保存过程。虽然FFPE是显微镜下染色组织分析的理想选择，但它会交联、破坏或去除组织中的蛋白质，使蛋白质含量的分析具有挑战性。迄今为止，报道的方法在过程和疗效方面差异很大。本教程介绍了一种源于系统测试和关键参数优化的策略，用于FFPE组织中蛋白质的可重复原位胰蛋白酶消化和随后的MALDI IMS分析。该方法描述了一种适用于几乎任何来源的FFPE组织的通用方法。
Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is a molecular imaging technology uniquely capable of untargeted measurement of proteins, lipids, and metabolites while retaining spatial information about their location in situ. This powerful combination of capabilities has the potential to bring a wealth of knowledge to the field of molecular histology. Translation of this innovative research tool into clinical laboratories requires the development of reliable sample preparation protocols for the analysis of proteins from formalin-fixed paraffin-embedded (FFPE) tissues, the standard preservation process in clinical pathology. Although ideal for stained tissue analysis by microscopy, the FFPE process cross-links, disrupts, or can remove proteins from the tissue, making analysis of the protein content challenging. To date, reported approaches differ widely in process and efficacy. This tutorial presents a strategy derived from systematic testing and optimization of key parameters, for reproducible in situ tryptic digestion of proteins in FFPE tissue and subsequent MALDI IMS analysis. The approach describes a generalized method for FFPE tissues originating from virtually any source.