To identification the immunization activation of recombinant Leptospira gene vaccine from many-siden MethodsThe guinea pigs were immunized with recombinant Leptospira gene vaccine [plasmid vector pT7-7 was negtive control ,inactivated whole cell vaccine (WCV) was positive control]. Then spleen cells were taken out. Particularity lymphocyte transformation test(LTT),IL-2 and IL-6 activation of these spleen cells were determined by MTT and 3H-TdR respectively. Results1)The Relative transformation index of gene vaccine group was significance higher than pT7-7 group (vaccin group: 2. 19±0. 18, pT7-7 group 1.42±0. 27 ( P＜0. 005 ); 2 ) the activation of IL- 2 and IL- 6 from recombinant gene vaccine group waw significance stronger than pT7-7 group (vaccine group IL-2:34. 8±3.11,IL-6:94. 6±6.03, pT7-7 group IL-2:20. 4±3. 05,IL-6: 67±6.28), (P＜0. 005). Conclusion1) The activation of Th1 and Th2 lymphocyte cell were increased in the guinea pigs with gene vaccine immunized. It suggested the recombinant Leptospira gene vaccine could elicit an extremely strong immunization effect of T-cell cooperate with B-cell and the gene vaccine was equal the WCV(P＞0. 05)but the gene vaccine sideeffect was small and the applying prospect was good.

AIM: To study the recombinant plasmid pDL121 and its expression product in E.coli. METHODS: pDL121 was analysed by using 6 different restriction endonucleases and dot blotting, and the isolated 23 kDa protein band was cut and injected twice into rabbits to raise anti - 23 kDa serum. RESULTS: The restriction map of pDL121 was quite different from other leptospiral genes reported and digoxin labeled recombinant DNA probe of pDL121 could detect the pathogenic leptospires, whereas, not the nonpathogenic leptospires. The anti - 23 kDa serum could recognize the sonicated antigen of L. interrogans serovar lai strain 017 and 23 kDa protein expressed in pDL121 and the titer of the antiserum were very high, approximately 1/12 800. Injection of the E. coli lysate of pDL121 with Freund's adjuvant into guinea pigs resulted in some protection of the animal against the challenge with strain 017. CONCLUSION: It indicated that 23 kDa protein had good imunogenicity and could serve as a candidate for protective antigen of L. interrogans and the inserted fragment of pDL121 could be a new gene .

AIM: To study the recombinant plasmid pDL121 and its expression product in E.coli. METHODS: pDL121 was analysed by using 6 different restriction endonucleases and dot blotting, and the isolated 23 kDa protein band was cut and injected twice into rabbits to raise anti-23 kDa serum. RESULTS: The restriction map of pDL121 was quite different from other leptospiral genes reported and digoxin labeled recombinant DNA probe of pDL121 could detect the pathogenic leptospires, whereas, not the nonpathogenic leptospires. The anti-23 kDa serum could recognize the sonicated antigen of L.interrogans serovar lai strain 017 and 23 kDa protein expressed in pDL121 and the titer of the antiserum were very high, approximately 1/12 800. Injection of the E.coli lysate of pDL121 with Freund's adjuvant into guinea pigs resulted in some protection of the animal against the challenge with strain 017. CONCLUSION: It indicated that 23 kDa protein had good imunogenicity and could serve as a candidate for protective antigen of L.interrogans and the inserted fragment of pDL121 could be a new gene .

Objective. To evaluate how the efficacy of DNA inocutation affects the ability to raise protective immunity against Leptospira.Methods. A pair of oligonucleotide primers were designed to amplify the endoflagellar gene of L. interrogans sensu stricto serovar lai. An approximately 840bp fragment was generated with PCR and inserted into VR1012, a plasmid DNA expression vector, after the fragment and VR1012 were digested respectively with EcoRV and Sal I. A recombinant plasmid designated as VR1012+flaB2 was obtained. The vector, VR1012 consits of a pUC18 backbone with the cytomegalovirus(CMV) IE1 enhancer, promoter, and intron A, transcription regulatory elements and the BGH polyadenylation sequences driving the expressing of leptospiral endoflagellar gene of L. interrogans sensu stricto serovar lai. Plasmid encoding leptospiral endoflagellin gene was injected into quadriceps of NZW rabbits.Results.This resulted in the generation of specific leptospiral antibody with high ELISA titer (1:32768) in the rabbits. Immuno/protection was performed in guinea pigs without adjuvant. The group"VR1012+flaB2" showed higher survival rate(90%,9/10 animals),compared with the group "VR1012 lack flaB2" and the group "normal saline".Conclusion.The technique of DNA vaccine has potential advantages over certain other vaccine preparation technologies. However whether DNA vaccine will be useful for vaccine development remains to be tested.