Western Blotting Protocol and its Troubleshooting

Abstract

Introduction: The detection of protein expression in tissues and nerve cells is necessary for the modern neuroscience research laboratory and laboratories making diagnostic approaches. To this point, we collected a comprehensive review article on steps and troubleshooting of Western blotting (WB). WB provides good qualitative results for determining a sample containing a target protein and is particularly useful for analyzing insoluble proteins. This process basically can be divided into five steps: (I) the protein extraction, (II) electrophoresis, (III) membrane transfer, (IV) immunodetection, and (V) visualization and analysis. These processes involve the transfer of protein patterns from gel to microporous membrane. Protein transfer with subsequent immunodetection is a great tool to identify a multitude of proteins, especially those proteins that are of low abundance. The efficient transfer of proteins from a gel to a solid membrane and specific immunodetection depend greatly on different parameters. Conclusion: For reducing the troubleshooting and enhancing awareness of the new developments in WB, this review summaries a representative of methods, reagents, and devices in the WB procedure.e.