Analysis of immune-reactive components of ayu serum using monoclonal antibody

本研究的目的是制备抗阿鱼血清成分，特别是主要免疫活性成分的单抗，以检测和定量测定其主要免疫活性成分。通过免疫印迹和以阿鱼血清致敏的聚苯乙烯小球，用过氧化物酶标记的抗小鼠免疫球蛋白，筛选和克隆已建立的小鼠骨髓瘤细胞和淋巴细胞的杂交瘤细胞株，反复进行制备单抗的试验。用鳗弧菌免疫的AYU血清和鳗弧菌脂多糖致敏的聚苯乙烯小球制备抗AYU单抗。对免疫和未免疫雏鸡的血清和肾、脾、肝组织匀浆进行检测。用孵化场的蛋和鱼苗进行检测，共获得12株单抗(“A”-“L”)。单抗“A”被认为是针对AYU抗体的。在血清、肾、脾、肝和鱼苗的匀浆中检测到与单抗“A”-“L”相对应的抗原“a”-‘1，而在虫卵中只检测到“b”-“e”和“1”，免疫后的阿育病毒浓度高于未免疫的阿育病毒，但其他抗原的浓度并没有增加。

The subject of the present study was to make monoclonal antibodies against serum components especially main immune reactive components of ayu, Plecoglossus altivelis, to detect and quantitatively determine those components.Trials to make monoclonal antibodies were done repeatedly by screening and cloning hybridomas of established mouse myeloma cells and lymphocytes obtained from the spleen of mice immunized with ayu serum by means of immune blotting and enzyme-linked immunosorbent assay (ELISA) with polystyrene beads sensitized with ayu serum using anti-mouse IgG labeled with peroxydase. To make anti-ayu antibody monoclonal antibody, ELISA using the serum of ayu immunized with Vibrio anguillarum and polystyrene beads sensitized with V.anguillarum lipopolysaccharide. Detection was done on the serum and homogenates of the kidney, spleen and liver of immunized and unimmunized ayu. Detection was dine also on eggs and fry of ayu obtained from a hatchery.Totally twelve monoclonal antibodies ("A"-"L") were obtained. Monoclonal antibody "A" was considered to be that against ayu antibody. Antigens "a"-'1", which corresponded to monoclonal antibodies "A"-"L", were detected in the serum and the homogenates of the kidney, spleen and liver and the fries but only "b"-"e" and "1" were detected in the eggs. Concentration of the "a" was higher in the immunized ayu than that in the unimmumized ayu, but the concentration of other antigens did not increasae by immunization of ayu.