[Efficient expression of exogenous human gp96 gene in tumor cells].
OBJECTIVE
Immunization of mice with preparations of heat shock protein(HSP) gp96 isolated from cancer cells has been shown to elicit specific protective cytotoxic T lymphocyte response against cells from which gp96 originate. This phenomenon exploits a new practicable pathway for cancer immunotherapy. But gp96 is generally expressed at low level in cells. Gp96 preparations from limited cells or tissue are difficult to meet the needs of study. So the current study aims to acquire minoclonal cell lines expressing gp96 at high level in order to prepare enough gp96 with high quality.


METHODS
The recombinant plasmid pcDNA-hgp96 of human gp96 cDNA was constructed by ligating the fragment of gp96 cDNA into the pcDNA3 plasmid, a eucaryotic expressing vector. Then the recombinant plasmid was transfected into Hela cells and the stable transfectants were selected with G418. The expression level of gp96 of the positive monoclonal cells was detected by Western blot analysis.


RESULTS
The recombinant expression plasmid of human gp96 cDNA was successfully constructed. The monoclonal cell lines with stable transfection were obtained. A monoclonal cell line expressing gp96 on high level was selected out.


CONCLUSIONS
The monoclonal cell line expressing gp96 at high level has been successfully established, which lays the groundwork for the study of its antitumor immunity.
Yue
Pei-bin
PB
National Center for Clinical Laboratory, Beijing Hospital, Ministry of Health, Beijing 100730, P. R. China.
Yang
Shu-de
SD
Huang
Chang-zhi
CZ
Zhao
Mei
M
Du
Fei
F
Lin
Liang
L
Yu
Quan
Q
Zhou
Qi-bing
QB
D000951:Antigens, Neoplasm / Q000096:biosynthesis / Q000235:genetics*; D018076:DNA, Complementary / Q000032:analysis; D006367:HeLa Cells; D006801:Humans; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis*; D014162:Transfection
D000951:Antigens, Neoplasm; D018076:DNA, Complementary; D011994:Recombinant Proteins; C052646:sarcoma glycoprotein gp96 rejection antigens