基于单克隆抗体的放射免疫分析法检测猪肾中的赭曲霉毒素A。
Detection of ochratoxin A in porcine kidneys by a monoclonal antibody-based radioimmunoassay.
作者信息
Rousseau D M, Candlish A G, Slegers G A, Van Peteghem C H, Stimson W H, Smith J E
出版信息
Appl Environ Microbiol. 1987 Mar;53(3):514-8. doi: 10.1128/aem.53.3.514-518.1987.
By using an indirect enzyme-linked immunosorbent assay, eight monoclonal antibodies (MAbs) were selected. Mice were immunized with ochratoxin A that was conjugated to bovine serum albumin. The hybridoma cell line designated 10G2 was grown in tissue culture and as an ascites tumor. The MAb was characterized to be specific to ochratoxin A and of the immunoglobulin G (IgG) class. Subsequently, the ascites fluid of this hybridoma was used in a competitive solid-phase IgG radioimmunoassay on protein A-Sepharose CL-4B, with [14C]ochratoxin A as tracer. Porcine kidneys were extracted with 0.5% phosphoric acid in chloroform. A two-step cleanup was achieved on a Sep-Pak C18 cartridge and a Sep-Pak silica cartridge. Radioimmunoassay with MAbs coupled to protein A-Sepharose CL-4B allowed the detection of ochratoxin A in porcine kidneys at a concentration as low as 0.2 ng/g.
通过使用间接酶联免疫吸附测定法，筛选出了8种单克隆抗体（MAb）。用与牛血清白蛋白偶联的赭曲霉毒素A免疫小鼠。命名为10G2的杂交瘤细胞系在组织培养中生长并形成腹水瘤。该单克隆抗体被鉴定为对赭曲霉毒素A具有特异性，且属于免疫球蛋白G（IgG）类。随后，将该杂交瘤的腹水溶液用于在蛋白A-琼脂糖凝胶CL-4B上进行的竞争性固相IgG放射免疫测定，以[14C]赭曲霉毒素A作为示踪剂。用0.5%磷酸的氯仿溶液提取猪肾。在Sep-Pak C18柱和Sep-Pak硅胶柱上进行两步净化。将单克隆抗体与蛋白A-琼脂糖凝胶CL-4B偶联进行放射免疫测定，能够检测到猪肾中低至0.2 ng/g浓度的赭曲霉毒素A。
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