Chapter 20 Detection of Flagellar Protein Kinases on Polyvinylidene Difluoride Membranes Following Sodium Dodecyl Sulfate—Polyacrylamide Gel Electrophoresis

摘要: Publisher Summary After electrophoretic separation, there is in situ renaturation of protein kinases polyacrylamide gels. Renaturation on polyvinylidene difluoride (PVDF) membranes enabled the detection up to 20 fibroblast culture cells, and has been used identify nuclear detect CaM kinase II purified samples or crude tissue homogenates. Protein are known be present cilia flagella many organisms involved control motility, mating, nondynein surface dynein force generation. This chapter describes a procedure for flagella. It demonstrated using demembranated ram sperm Chlamydomonas axonemes, but may applied other organisms. Ram axonemal active under reactivation promoting conditions discussed here. The method details solutions, preparation demembranation flagella, SDS-PAGE gradient gel electrophoresis, transfer proteins PVDF membrane subsequent renaturation. Unless retention plasma specifically desired, it best work with lanes blot show less background when used, bands appear sharper.