在Balb/c小鼠中开发抗人IgA单克隆抗体并进行特性鉴定。

Development and characterization of monoclonal antibodies against human IgA in Balb/c mice.

作者信息

Ezzatifar Fatemeh, Majidi Jafar, Baradaran Behzad, Maleki Leili Aghebati, Abdolalizadeh Jalal, Yousefi Mehdi

机构信息

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran Department of Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran.

Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran Department of Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran.

出版信息

Hum Antibodies. 2014;23(1-2):7-12. doi: 10.3233/HAB-140277.

DOI:10.3233/HAB-140277

PMID:25812697

Abstract

OBJECTIVE

Immunoglobulin A (IgA) is the second immunoglobulin in human serum. Monoclonal antibodies have many potential uses in diagnosis, treatment and purification. The conjugated monoclonal antibodies against human IgA are utilized in diagnostic kits.

METHODS

For production of monoclonal antibodies against human Immunoglobulin A, spleen cells of the immune mouse were fused with SP2/0 as myeloma cells and Poly Ethylene Glycol (PEG). Supernatant of hybridoma cells was screened to determine the antibody by ELISA. The appropriate clones were selected for limiting dilution (L.D). Ultimately, appropriate monoclone was injected intraperitoneally into the mouse that has been primed with Pristane.

RESULTS

In current study, 175 clones were obtained of which 5 clones had absorbance values of about 3 were selected for limiting dilution. Between these clones, 3-D5monoclone with IgG1 subclass was selected as appropriate one and it was reproduced in FCS free RPMI 1640. For large scale production in invivo, the appropriate clone was implanted in the peritoneum of the Balb/c mouse and its titer was determined, which showed 1/100,000 dilution for ascitic fluid, having no cross reactivity with IgM & IgG.

CONCLUSIONS

Monoclonal antibody was purified by chromatography, confirmed by SDS-PAGE and then conjugated with enzyme and used for diagnostic kits.

摘要

目的

免疫球蛋白A（IgA）是人类血清中的第二种免疫球蛋白。单克隆抗体在诊断、治疗和纯化方面有许多潜在用途。抗人IgA的偶联单克隆抗体用于诊断试剂盒。

方法

为制备抗人免疫球蛋白A的单克隆抗体，将免疫小鼠的脾细胞与SP2/0骨髓瘤细胞用聚乙二醇（PEG）进行融合。通过ELISA筛选杂交瘤细胞的上清液以确定抗体。选择合适的克隆进行有限稀释（L.D）。最终，将合适的单克隆细胞腹腔注射到已用 pristane 预处理的小鼠体内。

结果

在当前研究中，获得了175个克隆，其中选择了5个吸光度值约为3的克隆进行有限稀释。在这些克隆中，选择具有IgG1亚类的3-D5单克隆作为合适的克隆，并在无胎牛血清的RPMI 1640中进行增殖。为了在体内大规模生产，将合适的克隆植入Balb/c小鼠的腹腔并测定其效价，腹水效价显示为1/100,000稀释，与IgM和IgG无交叉反应。