Severe acute respiratory syndrome (SARS) is a serious infectious threat to public health. To create a novel trial vaccine and evaluate its potency, we attempted to generate a SARS inactivated vaccine using SARS coronavirus (SARS-CoV) strain F69 treated with formaldehyde and mixed with Al(OH)3. Three doses of the vaccine were used to challenge three groups ofBALB/c mice.We found that the mice exhibited specific IgM on day 4 and IgG on day 8. The peak titers of IgG were at day 47 in low-dose group 1∶19,200) and high-dose group (1∶38,400) whereas in middle-dose group (1∶19,200), the peak was at day 40. On day 63, the IgG levels reached a plateau. Neutralization assay demonstrated that the antisera could protect Vero-E6 cells from SARS-CoVs infection. Analysis of the antibody specificity revealed that the mouse antisera contained a mixture of antibodies specifically against the structure proteins of SARS-CoV. Furthermore, the mouse antisera conferred higher amount of antibodies against protein N, polypeptide S4 and S2 than those of proteins M and 3CL. These findings suggest that the inactivated SARS-CoV could preserve its antigenicity and the inactivated vaccine can stimulate mice to produce high levels of ntibodies with neutralization activity. Results also suggest that polypeptides originating from protein N or S might be a potential target for the generation of a recombinant SARS vaccine. © 2004 Elsevier B.V. All rights reserved.

Background The etiologic agent of severe acute re spiratory syndrome (SARS) ha s been confirmed to be a novel coronavirus (CoV), namely SARS2CoV1 Developing safe and effective SARS2CoV vaccine s is e ssential for us to prevent the po ssible reemergence of it s epidemic1 Previous experience s indicate that inactivated vaccine is conventional and more hopeful to be succe ssfully developed1 Immunogenicity evaluation of an experimental inactivated SARS2CoV vaccine in rabbit s wa s conducted and reported in this paper1 Methods The large2scale cultured SARS2CoV F69 strain wa s inactivated with 014 % formaldehyde and purified, then used a s the immunogen combined with Freund’s adjuvant1 Eight adult New Zealand rabbit s were immunized four time s with this experimental inactivated vaccine1 Twelve set s of rabbit serum were sampled from the third day to the seventy2fourth day after the first vaccination1 The titers of specific anti-SARS2CoV IgG antibody were determined by indirect enzyme2linked immuno sorbent a ssay, and the neutralizing antibody titers were detected with micro2cytopathic effect neutralization te st1 Results Rapid and potent humoral immune re sponse s were induced by the inactivated SARS2CoV vaccine in all the eight te st rabbit s1 Titers of both specific IgG antibody and neutralizing antibody peaked at about six weeks after first vaccination, with the maximum value of 1∶81 920 and 1∶20 480,re spectively1 After that, serum antibody levels remained at a plateau or had a slight decrea se, though two boo sters were given in the succedent 4 to 5 weeks1 Cro ss neutralization re sponse existed between SARS2CoV F69 strain and Z22Y3 strain1 Co ncl us i o ns The inactivated SARS2CoV vaccine made from F69 strain owns strong immunogenicity, and the cro ss neutralization re sponse between the two different SARS2CoV strains give s a hint of the similar neutralizing epitope s, which provide stable ba se s for the development of inactivated SARS2 CoV vaccine s1

ficial multidomain protein, was selected as the model molecule of complex protein with 2 disulfide bonds. A BbFGF-producing plasmid, pJN-BbFGF, and a HBscFv producing-plasmid, pQE-HBscFv, were constructed and transformed into E. coli strains BL21(DE3) and M15[pREP4] respectively. At the same time, both plasmids were transformed into a reductase-deficient host strain, E. coli Origami(DE3). The 4 recombinant E. coli strains were cultured and the target proteins were purified. Solubility and bioactivity of recombinant BbFGF and HBscFv produced in different host strains were analyzed and compared respectively. RESULTS: All recombinant E. coli strains could efficiently produce target proteins. The level of BbFGF in BL21(DE3) was 15-23% of the total protein, and was 5-10% in Origami (DE3). In addition, 65% of the BbFGF produced in BL21(DE3) formed into inclusion body in the cytoplasm, and all the target proteins became soluble in Origami (DE3). The bioactivity of BbFGF purified from Origami(DE3) was higher than its counterpart from BL21(DE3). The ED50 of BbFGF from Origami(DE3) and BL21(DE3) was 1.6μg/L and 2.2μg/L, respectively. Both HBscFv formed into inclusion body in the cytoplasm of M15[pQE-HBscFv] or Origami[pQE-HBscFv]. But the supernatant of Origami [pQE-HBscFv] lysate displayed weak bioactivity and its counterpart from M15[pQE-HBscFv] did not display any bioactivity. The soluble HBscFv in Origami[pQE-HBscFv] was purified to be 1-2 mg/L and its affinity constant was determined to be 2.62×107 mol/L. The yield of native HBscFv refolded from inclusion body in M15[pQE-HBscFv] was 30-35 mg/L and the affinity constant was 1.98×107 mol/L. There was no significant difference between the bioactivity of HBscFvs refolded from the inclusion bodies produced in different host strains. CONCLUSION: Modification of the redox environment of E. coli cytoplasm can significantly improve the folding of recombinant disulfide-bonded proteins produced in it.

目的：研究鲨鱼口服液对15名2型糖尿病病人短期病情控制、症状影响。方法：采用单盲、交叉对照等方法，对鲨鱼口服液受试者的血、尿常规，生化指标、血糖及糖化血清蛋白，以及临床症状等指标进行考察，观察时间为20 d。结果：患者在服用鲨鱼口服液可能改善糖尿病的症状，试验前10.7±6.4分，服用口服液后降为5.8±5.3，与安慰剂对比（P<0.05）。同时该试验品对糖尿病患者的肝、肾、心脏功能及血液学功能等无明显不良影响。结论：鲨鱼口服液在2型糖.尿病患者短期应用安全、可能改善症状、应用简便，但对血糖的影响仍需要长期观察。