Preparation and characterization of monoclonal antibody against melatonin.
Anti-melatonin monoclonal antibodies (MAb) were prepared following coupling melatonin to bovine serum albumin (BSA) by Mannich reaction. Balb/c mice were immunized via injection of the melatonin-BSA intraperitonally. The spleen cells producing high titer of antibody were fused with myeloma cells of SP2/0 origin. After two limiting dilutions, two stable clones (AS-H10 and AS-D26) exhibiting best properties were selected for further studies. The class and subclass of two MAbs were found to be IgG(1) and IgG(2a) with lambda and kappa light chains, respectively. Antibodies secreted by these two clones showed high affinity of about 10(9)M(1). Study of the specificity criteria showed that these clones had no cross reactivity with indolic, aromatic, and imidazole ring-containing compounds, and had high specificity towards melatonin. The calibration curve was constructed with a sensitivity range of 10 ng/mL to 10 microg/mL. In conclusion, these MAbs may be useful for immunoassay of melatonin.
Soukhtanloo
Mohammad
M
Department of Clinical Biochemistry, School of Medicine, Medical Sciences/University of Tehran, Iran.
Ansari
Mohammad
M
Paknejad
Maliheh
M
Parizadeh
Mohammad Reza
MR
Rasaee
Mohammad Javad
MJ
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002138:Calibration; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D006825:Hybridomas / Q000378:metabolism; D007074:Immunoglobulin G / Q000737:chemistry; D008550:Melatonin / Q000737:chemistry*; D051379:Mice; D008807:Mice, Inbred BALB C; D008956:Models, Chemical; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013154:Spleen / Q000378:metabolism
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D012710:Serum Albumin, Bovine; D008550:Melatonin