Production, characterization and applications of mouse anti-grass carp (Ctenopharyngodon idellus) growth hormone monoclonal antibodies.
Mouse anti-grass carp growth hormone (gcGH) monoclonal antibody (MAb) secretors were produced by PEG-mediated fusion of NS-1 myeloma cells and splenic B-lymphocytes of gcGH hyper-immunized mice. Positive secretors were screened by direct ELISA and cloned by limiting dilution. Three positive secretors, 21D3, 22G5 and 23B3, were obtained in a single fusion trial. Anti-gcGH MAbs were produced by growing hybridomas in the peritoneal cavity of pristane-primed mouse. The three MAbs were isotyped to be IgG2a, IgG2b and IgM, respectively. IgG MAbs were purified from ascitic fluid by Hitrap protein G column and IgM MAb was purified by gel filtration chromatography. The purified MAbs were highly specific and had moderate binding affinity. The MAbs were successfully used for the purification of native gcGH from mature grass carp pituitary extract by one-step immunoaffinity chromatography, for the quantification of gcGH by competitive sandwich ELISA, and for the probing of somatotropes in grass carp pituitary by immunohistochemistry.
Yiu-Kwong Leung
Michael
M
Department of Biochemistry, BMSB, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong, P.R. China.
Kwok-Keung Ho
Walter
W
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D001202:Ascitic Fluid; D001402:B-Lymphocytes / Q000378:metabolism; D002347:Carps / Q000276:immunology*; D002459:Cell Fusion; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D013006:Growth Hormone / Q000276:immunology*; D006825:Hybridomas; D007114:Immunization; D007074:Immunoglobulin G / Q000302:isolation & purification; D007075:Immunoglobulin M / Q000302:isolation & purification; D051379:Mice; D008807:Mice, Inbred BALB C; D009101:Multiple Myeloma / Q000378:metabolism; D010902:Pituitary Gland / Q000276:immunology / Q000378:metabolism; D013154:Spleen / Q000378:metabolism
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D013006:Growth Hormone