ABSTRACT
To isolate and culture the purified monoclonal neural stem cells from the cerebral cortex of new born mice, new-born mice cerebral cortex was isolated and dissociated to single-cell suspension by mechanical trituration. The dissociated single cells were cultured in serum-free medium. After the formation of neurospheres, single-cell clone culture was performed by limiting dilution and the proliferated single-cell clones were harvested for subculture. Immunocytochemistry was used to detect the specific marker of neuroepithelial stem cells (Nestin) of the primary and monoclonal neurospheres. In the differentiated cells we detected the specific antigen of NF-200 and GFAP. Our results showed that the primary neurospheres expressed Nestin antigen positively. By limiting dilution, we cultured the cell lines from single-cell clone and the monoclonal neurospheres expressed Nestin and had capabilities of self-renewal, proliferation and the potentiality of differentiation into neurons and glial cells. It is concluded that monoclonal neural stem cells which have the ability of proliferation and multi-directional differentiation can be isolated and cultured from the cerebral cortex of new-born mice by limiting dilution.
ABSTRACT
Objective:To construct a monoclone cell line expressing M-HBsAg.Methods:Transfected SP2/0 with pRc/CMV-S 2S by Calcium Phosphate Method,selected the cells by G418,detected HBsAg by Dot-blot、ELISA and Western-blot hybridization.Results:By selection of G418,G418 resistent cells were obtained;tests of Western-blot hybridization and ELISA demonstrated their expression of aimed protein.Conclusion:Monoclone Cell Lines of SP2/0-S 2S were obtained.