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how does charge-based separation work on Simple Western?
Simple Western charge-based assays take place in a capillary. Just load your samples and off to the races you go! Samples prepared in ampholytes and required reagents are loaded into an assay plate and placed in Peggy Sue or NanoPro 1000. The prepared sample is automatically loaded into the capillary and separated by charge according to their pI before the resolve proteins are immobilized to the capillary wall via a proprietary, photoactivated capture chemistry. Target proteins are identified using a primary antibody and detected using an HRP-conjugated secondary antibody and chemiluminescent substrate. The resulting chemiluminescent signal is detected and quantitated.
WHAT DOES THE CHARGE-BASED ASSAY DATA LOOK LIKE?
Simple Western charge-based assay data is processed automatically in Compass for SW software. Sample data is displayed as an electropherogram where it's easier to see minute changes. Quantitative results such as pI value, signal intensity (area), % area, and signal-to-noise for each immunodetected protein are presented in the results table automatically.
From Your Peers
Learn how researchers at Manchester University have developed. assays encompassing clinical samples from Chronic Myeloid Leukaemia (CML-SC), Chronic Lymphocytic Leukaemia (CLL), Non-Small Cell Lung Cancer (NSCLC; tissue & plasma) and Endometrial Cancer using ProteinSimple’s Simple Western charge assay. This and other work with Protein Simple defines the utility of the Simple Western charge assay in detailed mapping of signaling cascades in model systems and cellular organelles.