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[Establishment of eukaryotic cell line stably expressing soluble human leucocyte antigen G1 by nucleofection]
Menghua Zeng1, Chongyun Fang, Shusen Wang
1Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan Hubei, P R China.
Objective:
To establish a eukaryotic cell line that can express soluble human leucocyte antigen G1 (sHLA-G1) stably.
Methods:
The recombinant plasmid pcDNA3-sHLA-G1 is transfected by a novel nonviral, electroporation-based gene transfer method termed nucleofection into the host cell lymphoblastoid cell line (LCL) 721.221 which does not express any HLA-classical I molecules. After selection by G418, the cell line stably expressing sHLA-G1 is identified by RT-PCR and Dot-ELISA with HLA-G1 specific monoclonal antibody MEM-G/9.
Results:
The efficiency of transfection for LCL721. 221 is about 14% by nucleofection. The specific band for sHLA-G1 was found by RT-PCR assay from the transfections and the protein of sHLA-G1 in the supernatant of the transfections was detected by Dot-ELISA assay. Both confirmed that the eukaryotic cell line expressing sHLA-G1 has been established successfully at genic and proteinic levels.
Conclusion:
In this study, the eukaryotic cell line expressing sHLA-G1 have been established successfully by nucleofection.
