Lin C T, Chen L H, Chan T S
J Histochem Cytochem. 1983 Jul;31(7):920-6. doi: 10.1177/31.7.6854005.
The rabbit antiserum and mouse monoclonal hybridoma antibody against porcine cytosolic aspartate aminotransferase (c-AAT) (or cytosolic glutamic oxaloacetic transaminase (c-GOT)) were produced and compared for the localization of c-AAT in rat liver. An indirect immunocytochemical technique was performed using peroxidase-conjugated goat immunoglobulin (Ig) G anti-rabbit IgG and peroxidase-conjugated rabbit IgG anti-mouse IgG as the second antibody. Rats were perfused with paraformaldehyde-lysine-periodate fixative and the liver fragments were immersed in 4% paraformaldehyde and transferred to 10% dimethyl sulfoxide overnight and subjected to cryostat sectioning. The rabbit IgG antibody, 3 individual monoclonal antibodies, and a mixture of these 3 monoclonal antibodies were applied to the tissue sections, respectively, using the same concentration. Under the same experimental conditions, the c-AAT was localized in each individual hepatocyte by both monoclonal and polyclonal antibodies. However, a mixture of three monoclonal antibodies gave stronger staining than a single monoclonal antibody; although two antibodies yield more intense staining than just one, it was still less intense than for three. The conventional rabbit polyclonal antibody against c-AAT produced more reaction product than the combined three monoclonal antibodies. It is concluded that for immunocytochemical study, the use of a single monoclonal antibody is sensitive enough to localize its tissue antigen under the present experimental condition. To obtain a stronger reaction product, a combination of several monoclonal antibodies, at least three or more, may give better staining.
制备了抗猪胞质天冬氨酸氨基转移酶（c - AAT）（或胞质谷草转氨酶（c - GOT））的兔抗血清和小鼠单克隆杂交瘤抗体，并比较了它们在大鼠肝脏中对c - AAT的定位情况。使用过氧化物酶偶联的山羊免疫球蛋白（Ig）G抗兔IgG和过氧化物酶偶联的兔IgG抗小鼠IgG作为第二抗体，进行间接免疫细胞化学技术检测。用多聚甲醛 - 赖氨酸 - 高碘酸盐固定剂灌注大鼠，将肝组织碎片浸入4%多聚甲醛中，转移至10%二甲基亚砜中过夜，然后进行冰冻切片。分别以相同浓度将兔IgG抗体、3种单克隆抗体以及这3种单克隆抗体的混合物应用于组织切片。在相同实验条件下，单克隆抗体和多克隆抗体均能将c - AAT定位到每个肝细胞中。然而，三种单克隆抗体的混合物染色比单一单克隆抗体更强；虽然两种抗体的染色比一种抗体更强，但仍比三种抗体的染色弱。传统的抗c - AAT兔多克隆抗体产生的反应产物比三种单克隆抗体的混合物更多。得出结论，在本实验条件下，对于免疫细胞化学研究，使用单一单克隆抗体足以灵敏地定位其组织抗原。为了获得更强的反应产物，几种单克隆抗体（至少三种或更多）的组合可能会产生更好的染色效果。
