Reliability of G-Protein-Coupled Receptor AntibodiesAuthor(s): Tokuna Josti
Antibodies to 19 subtypes of 1 and -adrenoceptors, muscarinic, dopamine, and galanin receptors, as well as vanilloid (TRPV1) receptors, were shown to be non-selective. These findings show that for antibodies against G-protein-coupled and maybe other receptors, lack of selectivity appears to be the rule rather than the exception. As a result, the formerly widely used validation of such antibodies by the absence of staining in the presence of blocking peptide, i.e. the antigen against which the antibody was generated, is no longer sufficient to confirm specificity. We propose that receptor antibodies be validated using at least one of the following techniques like disappearance of staining in knock-out animals of the target receptor, reduction of staining upon knock-down approaches such as siRNA treatment, selectivity of staining in immunoblots or immunocytochemistry for the target receptor vs. related subtypes when expressed in the same cell line, and/or antibodies raised against multiple distinct epitopes of a receptor yielding venomous antibodies.