Department of Neurology and Institute of Translational Research, Rigshospitalet, Glostrup, Denmark.
Department of Neurology and Translational Research Center, Rigshospitalet, Glostrup, Denmark.
Methods Mol Biol. 2024;2821:237-248. doi: 10.1007/978-1-0716-3914-6_19.
Immunoblotting, also termed western blotting, is a powerful method for detection and characterization of proteins separated by various electrophoretic techniques. The combination of sodium dodecyl sulfate-poly acrylamide gel electrophoresis (SDS-PAGE), having high separating power, immunoblotting to synthetic membranes, and detection with highly specific peptide antibodies, is especially useful for studying individual proteins in relation to cellular processes, disease mechanisms, etc. Here, we describe a protocol for the sequential detection of various forms of an individual protein using peptide antibodies, exemplified by the characterization of antibody specificity for different forms of the protein calreticulin by double SDS-PAGE immunoblotting.
免疫印迹，也称为蛋白质印迹，是一种通过各种电泳技术分离蛋白质的检测和鉴定的强大方法。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）具有高分离能力，与合成膜结合进行免疫印迹，以及使用高度特异性的肽抗体进行检测，这种组合特别适用于研究与细胞过程、疾病机制等相关的单个蛋白质。在这里，我们描述了一种使用肽抗体顺序检测单个蛋白质的各种形式的方案，以双 SDS-PAGE 免疫印迹为例，用于鉴定钙网蛋白不同形式的抗体特异性。
