[Expression of midkine fusion protein and preparation and application of its monoclonal antibodies].

AIM: To express midkine (MK) protein, prepare its monoclonal antibody (mAb), and detect its expression in tumor cells.

METHODS

The human MK gene fragment was cloned from kidney cancer tissues by RT-PCR and inserted into the prokaryotic expression vector pMS-31b. The MS2-MK fusion protein was expressed in E.coli after temperature induction. The purified fusion protein was used to immunize BALB/c mice to prepare mAbs against MK by routine hybridoma technique. The Ig subclasses of the mAbs were identified by ELISA. The expression of MK in tumor cells was detected with the mAbs.

RESULTS

MK gene was cloned and MS2-MK fusion protein was expressed successfully. Two hybridoma cell lines secreting anti-MK mAbs were obtained. The Ig subclass of the 2 mAbs was IgG1 and IgG2a, respectively. MK was expressed in gastric carcinoma cell line MGC803 and gastric cancer tissue.

CONCLUSION

The MS2-MK fusion protein was expressed in E.coli and the mAbs against MK were pepared, which allow futher research on MK function.