A fully automated capillary western system for absolute quantitation of endogenous PKC proteins: An approach to correlate protein quantity with function.

31 Background: The human prostate cell line LNCaP and the human myelocytic leukemia cell line U937 differ dramatically in their responses to the two protein kinase C (PKC) targeted ligands phorbol 12-myristate 13-acetate (PMA) and bryostatin 1 and show complex differences in the patterns of transcriptional responses that they induce. Quantitation of relative abundance of individual PKC isoforms in the two cell lines may help to link the downstream effects of the two compounds to these isoforms. Methods: Simple Western is a capillary-based automated Western system recently developed by ProteinSimple. All steps following sample preparation are fully automated in the Simple Western system, including sample loading, size-based protein separation, immunoprobing, washing, detection and data analysis. Simple Western is gel-free and blot-free, uses less amount of samples, and produces highly quantitative, reproducible information that cannot be generated using regular Western assays. Using the Simple Western syst...