A comparative study of polyclonal and monoclonal antibodies for immunocytochemical localization of cytosolic aspartate aminotransferase in rat liver.
A comparative study of polyclonal and monoclonal antibodies for immunocytochemical localization of cytosolic aspartate aminotransferase in rat liver.
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多克隆和单克隆抗体对大鼠肝脏胞质天冬氨酸转氨酶免疫细胞化学定位的比较研究。
DOI：
10.1177/31.7.6854005
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
T. Chan
中科院分区：
文献类型：
--
作者：
C. Lin;L. H. Chen;T. Chan
制备了抗猪胞质天冬氨酸转氨酶（c-AAT）（或胞质谷氨酸草酰乙酸转氨酶（c-GOT））的兔抗血清和小鼠单克隆杂交瘤抗体，并比较了 c-AAT 在大鼠肝脏中的定位。使用过氧化物酶缀合的山羊免疫球蛋白 (Ig) G 抗兔 IgG 和过氧化物酶缀合的兔 IgG 抗小鼠 IgG 作为第二抗体进行间接免疫细胞化学技术。用多聚甲醛-赖氨酸-高碘酸盐固定剂灌注大鼠，将肝脏碎片浸入4%多聚甲醛中并转移至10%二甲亚砜过夜并进行低温切片。使用相同的浓度分别将兔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。在相同的实验条件下，通过单克隆和多克隆抗体将c-AAT定位于每个单独的肝细胞中。然而，三种单克隆抗体的混合物比单一单克隆抗体的染色更强；尽管两种抗体比一种抗体产生的染色强度更高，但仍不如三种抗体的强度。传统的抗 c-AAT 兔多克隆抗体比组合的三种单克隆抗体产生更多的反应产物。结论是，对于免疫细胞化学研究，使用单一单克隆抗体足够灵敏，可以在目前的实验条件下定位其组织抗原。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）的组合可以提供更好的染色。
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.