[Preparation, identification and preliminary application of the polyclonal antibody against gastric cancer-related protein GCRG224].
OBJECTIVE
To prepare the polyclonal antibody against gastric cancer-related protein GCRG224.


METHODS
The thioredoxin/GCRG224 fusion protein was expressed in E.coli. The polyclonal antibody against GCRG224 was obtained by immunizing a rabbit with the purified GCRG224 protein. The titer and specificity of the antibody were determined by ELISA and Western blot, respectively. The expression of GCRG224 in paraffin-embedded tissue sections from normal gastric mucosal tissues and advanced gastric cancer was determined using immunohistochemistry technique.


RESULTS
The thioredoxin/GCRG224 fusion protein with relative molecular mass of 16.8 kDa was over-expressed in E.coli. The purity of the expressed product directly purified from a denaturing polyacrylamide gel was about 100%. The polyclonal antibody against GCRG224 was prepared. ELISA detection proved the titer of antiserum against GCRG224 was about 1:256,000. Western blot analysis showed that the antiserum could bind to the expressed fusion protein specifically. GCRG224 was found to have higher expression in gastric cancer tissues than in normal ones by immunohistochemistry.


CONCLUSIONS
The successful preparation of the polyclonal antibody against GCRG224 lays a foundation for further study of the biological function and the possible role of GCRG224 in the development of gastric carcinoma.
Wu
Yin-qiao
YQ
Department of Gastroenterology, South Building, General Hospital of PLA, Beijing 100853, China. wuyq56@hotmail.com
Wang
Meng-wei
MW
Wu
Ben-yan
BY
Wang
Gang-shi
GS
You
Wei-di
WD
Wang
Wei-hua
WH
D000818:Animals; D000906:Antibodies / Q000032:analysis* / Q000276:immunology* / Q000302:isolation & purification; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008297:Male; D009363:Neoplasm Proteins / Q000235:genetics* / Q000276:immunology; D011817:Rabbits; D013274:Stomach Neoplasms / Q000737:chemistry / Q000235:genetics* / Q000276:immunology
D000906:Antibodies; C474288:GCRG224 protein, human; D009363:Neoplasm Proteins