Preparation of monoclonal antibody specific for rabbit lens epithelial cell
- VernacularTitle:抗兔晶状体上皮细胞单克隆抗体的制备及特异性研究
- Author:
Na CAI
;
Jinsong ZHANG
;
Furong LIU
;
Jindan SONG
- Publication Type:Journal Article
- Keywords:
rabbit lens epithelial cell;
hybridoma;
McAb;
immunofluorescence;
immunohisto chemistry SP
- From:
Recent Advances in Ophthalmology
2000;20(6):385-387
- CountryChina
- Language:Chinese
-
Abstract:
Objective To prevent the after cataract induced by lens epithelial cells proliferation from postoperative cataract, monoclonal antibody (McAb) specific for rabbit lens epithelial cell is made, it will be the carrier further to be conjugated with cytotoxin. The conjugations will inhibit lens epithelial cells growth and not damage the other tissues of eye. Thereby McAb is the experimental bases of preventing after cataract.Methods BALB/c mice were immunized by mixture of rabbit lens epithelial cells and Freund's adjuant. The immunized mouse spleen cells were fused with parental mouse myeloma cells (BALB/c SP2/0) using polyethylene glycol (PEG-4000). The fused cells were selectively cultured by hypoxanthine, aminopterin and thymidine (HAT) culture medium. The specificities of the supernatant from hybridomas were tested by indirect immunofluorescence and immunohistochemistry SP (Streptavidin peroxdase conjugated method). The positive hybridomas were further cloned three times by methylcellulose culture medium to ensure monoclonality. At last, the consensual reaction of McAb was tested on human eye tissues.Results Hybridomas were produced by fusion of spleen cells of immunized mice and mouse myoloma cells (SP2/0) with PEG-4000, and grown selectively in medium containing HAT after 16 days. Antibodies of the supernatant from hybridomas were tested on frozen sections of rabbit lens epithelial cell by indirect immunofluorescence. Only a positive clone secreted McAb against antigen of rabbit lens epithelial cell. The specificity of McAb was tested on paraffin sections of whole rabbit eye and whole human eye by immunohistochemistry SP. The results indicated that McAb was only positive to rabbit lens epithelial cell membrane and it was negative to the other tissues of rabbit or whole human eye tissues.Conclusions McAb specific for rabbit lens epithelial cell was manufactured successfully. The specificity of McAb was strong. There was no consensual reaction on human eye tissues. It might be the experimental bases of further targeting chemotherapy on after cataract.
