Fig. 2.
A schematic representation of whole-cell ELISA based assay for GPCR surface expression and endocytosis. HEK-293 cells expressing FLAG-tagged receptors are immobilized onto a 24-well cell culture plate. Incubation of cells with the HRP-coupled anti-FLAG M2 antibody leads to the binding of antibody to the FLAG epitope at the N-terminus of the receptor. Upon addition of TMB solution which contains a chromogenic substrate for HRP, a light blue color appears in the wells. The reaction is stopped by adding 1 M H2SO4 which results in the appearance of light yellow color. Absorbance of the solution is measured at 450 nm.