Institute of Medical Biochemistry and Molecular Biology, University Medicine Greifswald, 17475 Greifswald, Germany.
Institute of Anatomy and Cell Biology, University Medicine Greifswald, 17489 Greifswald, Germany.
Int J Mol Sci. 2023 Jul 11;24(14):11308. doi: 10.3390/ijms241411308.
Tissue sections, which are widely used in research and diagnostic laboratories and have already been examined by immunohistochemistry (IHC), may subsequently provide a resource for proteomic studies, even though only small amount of protein is available. Therefore, we established a workflow for tandem mass spectrometry-based protein profiling of IHC specimens and characterized defined brain area sections. We investigated the CA1 region of the hippocampus dissected from brain slices of adult C57BL/6J mice. The workflow contains detailed information on sample preparation from brain slices, including removal of antibodies and cover matrices, dissection of region(s) of interest, protein extraction and digestion, mass spectrometry measurement, and data analysis. The Gene Ontology (GO) knowledge base was used for further annotation. Literature searches and Gene Ontology annotation of the detected proteins verify the applicability of this method for global protein profiling using formalin-fixed and embedded material and previously used IHC slides.
组织切片在研究和诊断实验室中被广泛应用，且已经通过免疫组织化学（IHC）进行了检测，尽管可获得的蛋白质的量非常少，但它们仍可为蛋白质组学研究提供资源。因此，我们建立了一种基于串联质谱的 IHC 标本蛋白质分析工作流程，并对特定脑区切片进行了特征描述。我们研究了从成年 C57BL/6J 小鼠脑切片中分离的海马 CA1 区。该工作流程包含了从脑切片中进行样品制备的详细信息，包括去除抗体和覆盖基质、分离感兴趣的区域、蛋白质提取和消化、质谱测量和数据分析。还使用了基因本体论（GO）知识库进行进一步注释。对检测到的蛋白质的文献检索和基因本体论注释验证了该方法对于使用福尔马林固定和包埋材料以及先前使用的 IHC 载玻片进行全局蛋白质分析的适用性。
