Preparation of monoclonal antibody against LRR-WSC domain of polycystin-1 and distribution of polycystin-1 in kidney tissues and kidney cell lines
- VernacularTitle:多囊蛋白-1 LRR-WSC区单克隆抗体的制备及其在免疫组织化学诊断中的应用
- Author:
Haidan ZHAO
;
Changlin MEI
;
Xuefei SHEN
;
Tianmei SUN
;
Shuzhong ZHANG
;
Wenjing WANG
;
Yume WU
;
Ji SONG
- Publication Type:Journal Article
- Keywords:
Polycystic kidney;
Antibody, momoclonal;
Polycystin-1;
Immunohistochemistry
- From:
Chinese Journal of Laboratory Medicine
2003;0(08):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To prepare and identify monoclonal antibody against LRR-WSC domain of polycystin-1 and to investigate the distribution of polycystin-1 in kidney tissues and kidney cell lines. Methods BALB/c mice were immunized with fusion protein PC1-e of polycystin-1 LRR-WSC domain. The splenocytes were fused with myeloma cells by PEG 4000 and the hybridomas were selected in HAT medium. The hybridoma clones secreting antibodies against polycystin-1 LRR-WSC domain were detected by enzyme-linked immunosorbent assay ( ELISA) and cloned by limiting dilution. The specificity of anti-polycystin-1 LRR-WSC domain monoclonal antibody from hybridoma was verified by ELISA and Western blot. The distribution of polycystin-1 in tissues and cells was detected by immunohistochemical method. Results One cell line of hybridoma secreting monoclonal antibody against polycystin-1 was established. Western blot analysis showed that the monoclonal antibody reacted strongly and specifically to polycystin-1 LRR-WSC domain. Distribution of polycystin-1 in fetal kidney was localized in tubular epithelium. In normal adult kidney tissues, our study showed that polycystin-1 was mainly expressed in the medullary collecting ducts and distal convoluted tubules. Positive staining was also found in the majority of cyst-lining epithelial ceEs of cystic tissue from autosomal dominant polycystic kidney disease ( ADPKD) patients. Expressions of polycystin-1 were found in either ADPKD cyst-lining epithelia cell line and LLC-PK1, clearly plasma membrane and intracytoplasmic staining of polycystin-1 were observed. Conclusion Specific monoclonal antibody against polycystin-1 LRR-WSC domain were obtained. The antibody is important to researching the mechanism of ADPKD. The distribution of polycystin-1 in kidney tissues and cells show that polycystin-1 was important in tubular elongation and the maintenance of tubular architecture.
