A novel monoclonal antibody against human GRK6 antigen.
G protein-coupled receptor kinase 6 (GRK6) plays a universal role in receptor desensitization, by acting as a receptor-G protein interface, thereby affecting serine/threonine kinases. In this study, a 20-aa-long peptide of human GRK6 C-terminus domain was synthesized and covalently coupled to keyhole limpet hemocyanin (KLH). A mouse monoclonal antibody against human GRK6 (anti-GRK6 MAb) was successfully prepared through hybridoma technique by immunizing BALB/c mice with synthesized GRK6426-446-KLH peptides. A high specificity and affinity strain of hybridoma 5D12 were established. The titer of the purified anti-GRK6 MAb was 1.28 × 10(6) measured by indirect ELISA. Western blot and immunocytochemistry experiments were also applied to characterize the antibody specificity. Antibody absorption assays showed that the anti-GRK6 MAb can be blocked by GRK6426-446 peptides. These results indicated that the antibody could bind to GRK6 antigen specifically. This MAb provides valuable support for further studies on the functional properties of GRK6.
1 Department of Microbiology, West China School of Preclinical and Forensic Medicine, Sichuan University , Chengdu, China .
Qiang
Q
Zhang
Xiaojun
X
Huang
Jing
J
Yang
Mingyuan
M
Li
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000737:chemistry; D000918:Antibody Specificity; D000941:Antigens / Q000032:analysis* / Q000276:immunology; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D054768:G-Protein-Coupled Receptor Kinases / Q000032:analysis* / Q000276:immunology; D006241:Haptens / Q000737:chemistry / Q000276:immunology; D006433:Hemocyanins / Q000737:chemistry / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D007114:Immunization; D018796:Immunoconjugates / Q000008:administration & dosage / Q000737:chemistry / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D010455:Peptides / Q000008:administration & dosage / Q000138:chemical synthesis / Q000276:immunology; D017434:Protein Structure, Tertiary
D000911:Antibodies, Monoclonal; D000941:Antigens; D006241:Haptens; D018796:Immunoconjugates; D010455:Peptides; D006433:Hemocyanins; D054768:G-Protein-Coupled Receptor Kinases; C089176:G-protein-coupled receptor kinase 6; C032808:keyhole-limpet hemocyanin