恶性疟P190抗原保守区Ⅰ、Ⅴ基因的亚克隆及在大肠杆菌中高效表达

作者采用PCR方法克隆了我国海南省FCC1/HN株P190抗原两个保守区基因,分别定名为P190CRI和P190CRV。基因片段经纯化后连接到pUC18载体中进行DNA序列分析,结果显示:除了P190CRV中有5个碱基变换外,其余序列均与MAD20型序列一致。经序列分析的两个基因片段分别与pGEX-2T载体连接,经双酶切鉴定后转化感受态JM109(DE_3)大肠杆菌进行高效融合表达,并且用Sepharose 4B-谷胱甘肽层析柱进行亲和纯化,结果为:两个插入基因片段均得到高效融合表达,经一步亲和纯化后就取得高纯度的重组蛋白。     

Dendritic Cells Coinjected with Tumor Cells Treated with an Anticancer Drug to Induce Tumor Rejection

Purpose: We examined whether bone marrow-derived dendritic cells (DCs) could induce antitumor immunity when a chemotherapeutic drug was added. Methods: CT26 (a murine colon cancer cell line syngeneic with BALB/c) and CT26-bearing mice were treated with mitomycin C (MMC) intraperitoneally (i.p.). Next, mice immunized with a coinjection of DCs and MMC-treated CT26 (i.p.) were given an intradermal inoculation of CT26. Finally, CT26-bearing mice were treated with MMC (i.p.) with or without DCs, given peritumorally. Results: Although the inoculated tumor was not rejected in the control mice, CT26 was rejected in 50% of the mice injected with MMC alone. Apoptosis was observed in the MMC-treated CT26 cells in vitro and in vivo. Immunization with DCs and apoptotic CT26 cells, but not with apoptotic CT26 alone, gave protection against tumor challenge in 7 of 13 mice. A significantly higher level of cytotoxic T-cell activity and interferon-γ production was seen in the protected mice. When MMC (i.p.) treatment was followed by peritumoral DC injection in the CT26-bearing mice, remarkable therapeutic effects were observed. Conclusion: DCs can collaborate with chemotherapy-induced apoptotic tumor cells and elicit improved antitumor immunity, probably through the acquisition of tumor-associated antigens from apoptotic tumor cells. Received: January 7, 2002 / Accepted: September 3, 2002 Acknowledgments. We thank Dr. Kazuo Kinoshita for his useful advice on using flow cytometry. This research was partly supported by the Ministry of Education, Culture, Sports, Science and Technology (No. 11671160). Reprint requests to: S. Yamasaki     

小剂量卡介苗、丝裂霉素交替灌注预防浅表性膀胱癌术后复发

目的 观察小剂量卡介苗(BCG)、丝裂霉素C(MMC)交替灌注预防浅表性膀胱癌术后复发的疗效和安全性。方法 回顾性分析自2000年5月～2001年6月的45例浅表性膀胱癌患的临床资料，29例行膀胱部分切除术，16例行经尿道膀胱肿瘤电切术(TuRBt)，术后定期应用卡介苗60mg、丝列霉素C 10mg交替膀胱灌注，每周1次共6次，间歇3个月，再每月1次，持续12个月。其中Ta、T1期22例、T2期23例。结果 所有病人随访12～24个月，45例患中，共有2例复发，均为TURBt后T2病人，总有效率95．2％，无全身不良反应，仅2例应用BCG后出现轻度膀胱刺激症状。结论 小剂量MMC、BCG交替膀胱内灌注，预防浅表性膀胱癌术后复发，效果好、病人耐受性强、副作用小。     

紫外线减毒尾蚴活疫苗预防小鼠日本血吸虫感染有效免疫期的实验研究

以 400μW/(min·cm~2)剂量紫外线照射减毒日本血吸虫尾蚴, 1 次免疫小鼠(200条/只)可产生抗日本血吸虫感染的免疫能力。这种免疫力至少可维持7个月。1、3、5、7个月各组中,免疫组与攻击组虫荷均数比较,差异均有显著性意义。1、3、5和7个月所获减虫率分别为71.28％、69.63％、62.26％和46.16％,7个月与1、3、5个月比较,差异有显著性意义,提示7个月时小鼠抗日本血吸虫的免疫力可能有所下降。     

Comparative vaccination of cattle against Boophilus microplus with recombinant antigen Bm86 alone or in combination with recombinant Bm91

Cattle were vaccinated either with a single recombinant tick antigen, Bm86 or with a combination of two recombinant antigens, Bm86 and Bm91 from the tick Boophilus microplus . In three experiments, the responses of cattle to subsequent challenge with the tick were assessed. The addition of the Bm91 antigen enhanced the efficacy of the vaccination over that with Bm86 alone to a statistically significant degree. Moreover, co-vaccination with two antigens did not impair the response of cattle to the Bm86 antigen. Finally, responses of individual cattle to the two antigens were independent. All of these results may be relevant to the increase in efficacy expected from a dual antigen vaccine.     

江苏省肾综合征出血热单价灭活疫苗中期免疫效果观察

观察HFRS单价灭活疫苗中期免疫效果。方法：采用IFAT法及MCPENT法检测荧光抗体及中和抗体。结果：3个试区加强接种人数分别为4052、4407和6354人。加强后，Ⅰ型苗荧光抗体阳性率为72．73％，GMT为14.14；中和抗体阳性率为54．55％，GMT为6．67。D型苗荧光抗体阳性率为75.00％，GMT为11.85；中和抗体阳性率为60％，GMT为9．44。加强后1年，Ⅰ型苗年均保护率为65．52％，Ⅱ型苗则为94．24％。同期，Ⅰ型苗荧光抗体阳性率维持在40％，中和抗体维持在35．00％；Ⅱ型苗荧光抗体阳性率降至10．34％，中和抗体维持在53．85％。结论：两型疫苗均有较好的中期防病效果。     

Epitope mapping and biological function analysis of antibodies produced by immunization of mice with an inactivated Chinese isolate of severe acute respiratory syndrome-associated coronavirus (SARS-CoV)

Inactivated severe acute respiratory syndrome-associated coronavirus (SARS-CoV) has been tested as a candidate vaccine against the re-emergence of SARS. In order to understand the efficacy and safety of this approach, it is important to know the antibody specificities generated with inactivated SARS-CoV. In the current study, a panel of twelve monoclonal antibodies (mAbs) was established by immunizing Balb/c mice with the inactivated BJ01 strain of SARS-CoV isolated from the lung tissue of a SARS-infected Chinese patient. These mAbs could recognize SARS-CoV-infected cells by immunofluorescence analysis (IFA). Seven of them were mapped to the specific segments of recombinant spike (S) protein: six on S1 subunit (aa 12-798) and one on S2 subunit (aa 797-1192). High neutralizing titers against SARS-CoV were detected with two mAbs (1A5 and 2C5) targeting at a subdomain of S protein (aa 310-535), consistent with the previous report that this segment of S protein contains the major neutralizing domain. Some of these S-specific mAbs were able to recognize cleaved products of S protein in SARS-CoV-infected Vero E6 cells. None of the remaining five mAbs could recognize either of the recombinant S, N, M, or E antigens by ELISA. This study demonstrated that the inactivated SARS-CoV was able to preserve the immunogenicity of S protein including its major neutralizing domain. The relative ease with which these mAbs were generated against SARS-CoV virions further supports that subunit vaccination with S constructs may also be able to protect animals and perhaps humans. It is somewhat unexpected that no N-specific mAbs were identified albeit anti-N IgG was easily identified in SARS-CoV-infected patients. The availability of this panel of mAbs also provided potentially useful agents with applications in therapy, diagnosis, and basic research of SARS-CoV.     

Restriction endonuclease analysis of varicella-zoster vaccine virus and wild-type DNAs

The DNA from several clinical isolates of varicella-zoster virus (VZV) were compared with the DNA from the vaccine strain VZV using three restriction endonucleases: BamHI, BgII, and HpaI. When electrophoresed through an agarose gel, the vaccine DNA digestion pattern was significantly different from the digestion patterns of the wild-type DNAs. Variations in the digestion pattern of the separate clinical isolates were also observed.     

脂质体转染AFP基因构建树突状细胞肝癌瘤苗及体外活性检测

目的：探讨以ＡＦＰ为靶点，构建ＡＦＰ－ＤＣ肝癌瘤苗及其抗肝癌免疫治疗的可能性。方法：应用脂质体将ＡＦＰ基因转染体外培养的未成熟 ＢＭＤＣ，构建 ＡＦＰ－ＤＣ肝癌瘤苗，以流式细胞术、Ｗｅｓｔｅｒｎ ｂｌｏｔ、３Ｈ－ＴｄＲ法和 ＭＴＴ法等检测其免疫活性。结果：ＡＦＰ－ＤＣ瘤苗不仅能产生和分泌ＡＦＰ，而且能上调自身的Ｂ７分子和ＭＨＣ分子，明显刺激Ｔ细胞增殖及提高ＣＴＬ的杀伤作用。结论：提示肝癌相关基因ＡＦＰ可作为抗肝癌基因治疗的切入点。     

Sequence Analysis of the Washington/1964 Strain of Human Parainfluenza Virus Type 1 (HPIV1) and Recovery and Characterization of Wild-Type Recombinant HPIV1 Produced by Reverse Genetics

A complete consensus sequence was determined for the genomic RNA of human parainfluenza virus type 1 (HPIV1) strain Washington/20993/1964 (HPIV1 WASH/64), a clinical isolate that previously was shown to be virulent in adults. The sequence exhibited a high degree of relatedness to both Sendai virus, a PIV1 virus recovered from mice, and human PIV3 (HPIV3) with regard to cis-acting regulatory regions and protein-coding sequences. This consensus sequence was used to generate a full-length antigenomic cDNA and to recover a recombinant wild-type HPIV1 (rHPIV1). Interestingly, the rHPIV1 could be rescued from full-length antigenomic rHPIV1 cDNA using HPIV3 support plasmids, HPIV1 support plasmids, or a mixture thereof. The replication of rHPIV1 in vitro and in the respiratory tract of hamsters was similar to that of its biologically derived parent virus. The similar biological properties of rHPIV1 and HPIV1 WASH/64 in vitro and in vivo, together with the previous demonstration of the virulence of this specific isolate in humans, authenticates the rHPIV1 sequence as that of a wild-type virus. This rHPIV1 can now be used to study the biological properties of HPIV1 and as a substrate to introduce attenuating mutations for the generation of live-attenuated HPIV1 vaccine candidates.An erratum to this article can be found at