Challenges of Western Blotting
Sunday, September 18, 2022
10:56 AM
Western blotting (WB) is a time consuming multi step process that can lend itself to errors in each step of the process; this can lead to many challenges. It would be safe to say that there is a lack of any consensus on what constitutes reproducibility. Not every researcher or lab technician has the same skill set which can add an additional layer of variability in the process. Primary antibody validation is critical to any WB protocol. Numerous examples of problems associated with improperly validated antibodies exist. Validation involves determining the antibodies specificity for its target and independent of lot-to-lot variations. Imaging after spending multiple years on a project only to find out that the antibody was binding to a protein other than the protein of interest. Unfortunately, there is no independent validation data for antibodies available. Variations in WB equipment used, experimental temperature, buffers used or the concentrations of antibodies used may result in different results from what is provided by the antibody manufacturer. A robotic WB system could eliminate errors in reproducibility leading to more reliable publication quality results. The blot processing steps are tedious, time consuming and potentially error prone. The BlotCycler touch or Mini (Precision Biosystems) performs complete automation of all membrane processing steps; these steps done manually are tedious and time consuming. Precision BlotCyclers allow for the recycling of primary antibody, increase throughput, lower antibody volume, and the benefit of running in the cold room. The result is low background, improved sensitivity and better time management.
