[Prokaryotic expression of human calcyclin-binding protein and preparation of mouse polyclonal antibody against hCacyBP].
OBJECTIVE
To express hCacyBP in E.coli and prepare mouse polyclonal antibody against hCacyBP so as to study tissue distribution of hCacyBP and evaluate its role during formation of multidrug resistance to gastric cancer.


METHODS
hCacyBP gene was subcloned into an expression vector pET28a, and then the recombinant vector was transformed into E. coli BL21. The recombinant protein was expressed in BL21 under IPTG induction. Using purified hCacyBP as immunogen, mouse polyclonal antibody against hCacyBP was prepared. hCacyBP was detected by Western blot in 10 kinds of rabbit tissues. Expression and distribution of hCacyBP in SGC7901/VCR and SGC7901 cells were detected by Western blot and immunohistochemical staining.


RESULTS
hCacyBP was successfully expressed in E. coli. The Western blot analysis showed that hCacyBP was expressed in all 10 kinds of rabbit tissues, but expression of brain and liver tissues were higher as compared with other tissues. Expression and distribution of hCacyBP in both SGC7901/VCR and SGC7901 cells had no significant difference.


CONCLUSIONS
CacyBP expressed widespreadly in varied tissues. Polyclonal antibody against hCacyBP that we prepared has high specificity, which provides a powerful tool for studying the function of hCacyBP.
Cheng
Yi
Y
Institute of Digestive Disease, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Yan
Peng-fei
PF
Qiao
Tai-dong
TD
Fan
Dai-ming
DM
D000818:Animals; D000918:Antibody Specificity; D001921:Brain / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D045744:Cell Line, Tumor; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D006801:Humans; D007106:Immune Sera / Q000096:biosynthesis* / Q000276:immunology; D008099:Liver / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D010957:Plasmids; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D013274:Stomach Neoplasms / Q000378:metabolism* / Q000473:pathology; D014169:Transformation, Bacterial
C491778:CACYBP protein, human; D002135:Calcium-Binding Proteins; D007106:Immune Sera; D011994:Recombinant Proteins