Abstract: To develop a soluble B lymphocyte stimulating factor (sBLyS) detection assay, the sBLyS encoding gene was cloned in this study, and sBLyS recombinant protein was expressed by using Escherichia coli BL21(DE3). The recombinant protein was purified by nickel column affinity chromatography. BALB/c mice were immunized with purified sBLyS recombinant protein, and cell lines secreting anti-BLyS monoclonal antibody (mAb) were obtained by using cell fusion technology. The mAb was purified by using n-Caprylic acid combined ammonium sulfate precipitation assay, and Western Blot and ELISA were used to analyze the activity and titer of the mAb. The unlabelled mAb and the HRP labeled mAb was used as the capture antibody, and the detection antibody, respectively, and then the double antibody sandwich ELISA test were performed to screen the optimal combination. The results showed that the sBLyS recombinant protein was expressed and purified. Four cell lines secreting the sBLyS monoclonal antibody hybridoma were obtained (named 2E4, 2H8, 4C5 and 4C12) with the titer of 9.6， 10.8, 20.4 and 26.6ng/mL, respectively. Western Blot results showed that the purified monoclonal antibodies had sBLyS protein binding activity. Double antibody sandwich ELISA showed that the combination of 2H8 and HRP-4C5 could be used to detect sBLyS, and the testing linearity ranged from 0.4ng/mL to 1.6ng/mL (R2=0.995).

Key words: autoimmunity disease, B lymphocyte stimulating factor, prokaryotic expression, monoclonal antibody