An On-Cell Western is a cell-based assay that enables quantitative monitoring of cell surface protein expression.
The On-Cell Western can be used to:
In standard In-Cell Western™ assays, cell membranes are permeabilized so that antibodies can reach antigens inside the cell.
With On-Cell Western assays, unpermeabilized cells are stained with antibodies against extracellular protein domains, so only cell surface antigens are detected.
Miller et al. used the assay to study internalization and recycling of CB1 (cannabinoid receptor), a G protein-coupled receptor, after treatment with receptor agonists and cycloheximide. The antibodies used were targeted against specific extracellular or intracellular domains of CB1. The observed time course of receptor internalization was consistent with previous confocal microscopy studies.
Delisle et al. adapted the assay to quantitatively measure plasmalemmal expression of hERG in live cells. The data correlated well with Western blot and whole-cell patch-clamp analyses.
See some published examples of On-Cell WesternsView publications