Establishment of Mouse CD40L Stably Expressed NIH3T3 Cell Lines and Its Application in the Culture and Activation of B Lymphocytes
LANG Qiaoli , HE Qilin, HUANG Nan, YU Lin, YANG Xi*
Recombinant plasmid pcDNA3.1-mCD40L was constructed and transfected into NIH3T3 cells to express mouse CD40L. The transfected cells were selected with G418 to obtain stably transfected cell lines. The RNA of stable transfected cells was isolated and the mRNA expression level of Neo gene was detected by RT-PCR. The expression level of CD40L protein in the culture supernatant of transfected cells was detected by ELISA analysis. RT-PCR revealed that Neo gene could be expressed in the transfected cells. ELISA results showed that the concentration of CD40L in the supernatant of the stable transfected cell line NIH3T3- mCD40L was 1.286 ng/mL. Further activity studies showed that the addition of NIH3T3- mCD40L cells, IL-2 and IL-21 could successfully culture and stimulate B cells to produce specific antibodies. NIH3T3-mCD40L stable cell lines was successfully obtained. This study lays a good foundation for generation of monoclonal antibody from B lymphocytes.
CN
EN