OBJECTIVETo discuss the optimal immunization dose by observing the immunoprotective effects of different doses of recombinant Schistosoma japonicum (Chinese strain) signaling protein 14-3-3 (rSj14-3-3).

METHODSSj14-3-3 gene was amplified by reverse transcriptase PCR (RT-PCR), subcloned into prokaryotic expression vector pET28a, then transformed into E.coli to express by inducing. Purified rSj14-3-3 was prepared through SDS polyacrylamide gel electrophoresis (SDS-PAGE), electroelution, dialysis, then BALB/c mice were divided into 5 groups and immunized in rSj14-3-3 protein followed by challenging infection (the 1st, 2nd, and 3rd groups were immunized in 50 microg, 100 microg and 300 microg antigen, respectively. The 4th, 5th groups were immunized in Freund's adjuvant and normal saline controls). After 6 weeks of challenging infection, the mice were killed and the worm and egg reduction rates were calculated. And the mice sera in different time were taken to examine the specific anti-Sj14-3-3 IgG.

RESULTSrSj14-3-3 protein was expressed successfully. After immunizing and challenging, worm reduction was found to be 28.20% in the 1st group, 43.10% in the 2nd group, 40.00% in the 3rd group, respectively. Number of eggs in liver tissue was reduced by 41.80%, 57.50%, 55.70%, respectively. Compared the results of the tested groups to the controls, the differences were of significance by t-test (worm reduction rate: t = 6.8 in the 1st group, t = 8.7 in the 2nd group, t = 7.3 in the 3rd group, P < 0.01 in all tested groups. Egg reduction rate at the group's number above: t = 11.23, t = 11.54, t = 7.99, P < 0.01 in all tested groups). As compared the results between the tested groups by chi(2), the differences were of significance between the 1st and the 2nd groups (worm reduction rate: chi(2) = 8.96, P < 0.05; egg reduction rate: chi(2) = 15.69, P < 0.05), between the 1st and the 3rd groups, the differences were also of significance (worm reduction rate: chi(2) = 6.52, P < 0.05; egg reduction rate: chi(2) = 12.52, P < 0.05). The difference was not of significance between the 2nd and the 3rd groups (worm reduction rate: chi(2) = 1.20, P > 0.05; egg reduction rate: chi(2) = 0.93, P > 0.05). In all tested groups, total anti-Sj14-3-3 specific IgG rose markedly. IgG(1) and IgG(2a) subtypes were high, but IgG(2b) and IgG(3) were near the background in four subtypes tested.

CONCLUSIONImmunoprotection of rSj14-3-3 should have some relations with immunization dose, and the protection obtained from immunizing mice by using 100 microg antigen was the best.