Should we be cautious on the use of commercially available antibodies to dopamine receptors?
Evidence indicate that it is difficult to obtain specific antibodies to G protein-coupled receptors and different technical difficulties may allow the generation of antibodies that lack specificity. We conducted experiments to validate the specificity of commercially available antibodies raised against dopamine (DA) receptors hD(1), hD(4), and hD(5) using a transfection approach: we studied whether, in HEK 293 cells selectively transfected with the various cloned subtypes, each antibody generates bands only in cells expressing its cognate receptor but not in those expressing the other DA receptors. Our results demonstrated that hD(1) and hD(4) receptor antibodies recognize not only their respective epitope, but also other DA receptor subtypes, while for the hD(5) receptor detection, we observed a signal only in the lane loaded with hD(5)-transfected HEK 293 cells, although with a lack of purity. Therefore, we recommend caution on the use of commercially available DA receptor antibodies.
Bodei
Serena
S
Section of Pharmacology, Department of Biomedical Sciences and Biotechnologies, University of Brescia Medical School, V.le Europa 11, 25124, Brescia, Italy.
Arrighi
Nicola
N
Spano
PierFranco
P
Sigala
Sandra
S
D000906:Antibodies / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology*; D015153:Blotting, Western; D002460:Cell Line; D006801:Humans; D011954:Receptors, Dopamine / Q000032:analysis* / Q000235:genetics / Q000276:immunology*; D017447:Receptors, Dopamine D1 / Q000032:analysis / Q000235:genetics / Q000276:immunology; D017448:Receptors, Dopamine D2 / Q000032:analysis / Q000235:genetics / Q000276:immunology; D050637:Receptors, Dopamine D3 / Q000032:analysis / Q000235:genetics / Q000276:immunology; D050638:Receptors, Dopamine D4 / Q000032:analysis / Q000235:genetics / Q000276:immunology; D050641:Receptors, Dopamine D5 / Q000032:analysis / Q000235:genetics / Q000276:immunology; D014162:Transfection
D000906:Antibodies; D011954:Receptors, Dopamine; D017447:Receptors, Dopamine D1; D017448:Receptors, Dopamine D2; D050637:Receptors, Dopamine D3; C113776:dopamine D2L receptor; D050638:Receptors, Dopamine D4; D050641:Receptors, Dopamine D5