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Expression of the extracellular domain of human CD7 in Escherichia coli

Immunological Journal.2001;(2):105-106,109. doi:10.3969/j.issn.1000-8861.2001.02.008

Objective　To get the antigen of CD7.Methods　 The extracellular domain of human CD7 was cloned from a plasmid containing the full length of human CD7 cDNA and expressed in pinpoint-xa3 prokaryotic system. Results　 Analysis with SDS-PAGE and Western-blotting showed that the expressed protein could bind to the anti-CD7 mAb specifically and is about 30 000 u in molecular weight. Conclusion　 These results paved the way for preparing anti-CD7 engineering antibody.

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Synthesis and identification of methylparaoxon artificial antigen

Weiguo ZHANG ; Zhaoyang MA

Immunological Journal.2001;(2):144-145,154. doi:10.3969/j.issn.1000-8861.2001.02.021

Objective　To synthesize the artificial antigen methylparaoxon （M1600）.Methods　Methylparaoxon was reduced into amino-methylparaoxon by using acetic acid-zinc power-hydrochloric acid. Artificial antigens M1600-BSA， M1600-TTH　were synthesized by conjugating amino-methylparaoxon to bovine serum albumin （BSA）and tachypleus tridentatus hemocyanin （TTH）directly after diazotization.Results　 Rabbits had been immunized with M1600-BSA for 10 weeks， and the high titer and high specificity antiserum from those rabbits was testified by doubled agar gel diffusion and indirect ELISA．Conclusion　An artificial antigen was obtained successfully and this made it possible to establish the immunoassay of M1600.

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Study of activation of human T cells with superantigen SEA

Guilian XU ; Xihua ZHU ; Jin YANG

Immunological Journal.2001;(2):85-87,104. doi:10.3969/j.issn.1000-8861.2001.02.002

Objective　To investigate the activation of human T cells with superantigen SEA. Methods　T cells proliferation, IL-2 production, DNA synthesis, cell phenotype and apoptosis induced by the SEA in the first or restimulation were detected. Results　 It was shown that 100 ng/mL was the optimal activation concentration, IL-2 production arrived at top level in the third day, SEA activated CD4+ and CD8+T with the same degree, T cells start apoptosis in response to SEA restimulation within 24 hours and apoptosis disappeared through addition of rhIL-2. Conclusions　 There were correlation between activation and SEA concentration or stimulation period； SEA activated both CD4+ and CD8+T cells without change the cell phenotype.

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Expression and relationship between HDAg and bcl-2／bax in liver tissue of the patients with hepatitis D

Xiaohong GU ; Qifen LI ; Yuming WANG

Immunological Journal.2001;(2):128-131. doi:10.3969/j.issn.1000-8861.2001.02.016

Objective　To study the role of bcl-2 and bax in the pathogenesis of hepatitis D. Methods　Expression of HDAg， bcl-2 and bax in liver specimens of 79 patients with hepatitis D were studied by immunohistochemistry technique. Meanwhile， the relationship between expression of HDAg and that of bcl-2／bax in liver specimens of patients were studied by double labelling technique and serial sections.Results　bcl-2 was mainly expressed in the cytoplasm of hepatocytes， bax mainly in the cytoplasm of hepatocytes and partly in the nucleus of hepatocytes， and HDAg mainly in the nucleus of hepatocytes. Most of HDAg and bax positive cells were distributed among infiltrating lymphocytes at the periportal region especially at the advancing edges of areas of “piecemeal necrosis”. Most of hepatocytes of bax positive was found to locate near to positive cells of HDAg， and there were positive correlations between degrees of bax expression and HDAg expression（P＜0.05）. Conclusions　The distribution and the expression of bax and HDAg are significantly correlated with the activity of inflammation and the severity of the liver damage， and HDV infection may induce the expression of bax in the hepatocytes，and hepatocyte apoptosis mediated by bcl-2／bax may play an important role in the liver cell injury by HDV infection.

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Expression of IL-18 in the peripheral blood mononuclear cells of chronic hepatitis B patients

Weiqun WEN ; Lian ZHANG ; Zhangyong HU ; Mingxia ZHANG ; Hong XIAO ; Yi QIAN

Immunological Journal.2001;(2):125-127. doi:10.3969/j.issn.1000-8861.2001.02.015

Objective　 To investigated the possible roles of IL-18 in HBV infection, and the expression of IL-18 in the peripheral blood mononuclear cells (PBMC) of chronic hepatitis B. Methods 　In 15 cases of asymptomatic carriers, 30 cases of active and remissive phases of chronic hepatitis and 10 healthy individuals (as normal controls), IL-18 expression in PBMC were quantitatively analyzed with flow cytometric immunological method. The PBMC were separated routinely and stimulated with LPS (SIGMA) and Monensin (SIGMA) for 6 hours. Then the cells were harvested and fixed by PBS/4% paraformaldehyde. The treated cells were stored in liquid nitrogen for detection later. After immunologic stain, the expression of IL-18 in PBMC were examined by flow cytometry. Results　 ① IL-18 was the lowest in asymptomatic carriers. It was fewer in remissive phase than in normal controls (P＜0.01).There was no difference between active phase and normal controls(P=0.25).②Within the group of active phase, IL-18 was significantly different between each grade of inflammatory activity(P＜0.01),and correlated with serum ALT positively(r=0.63,P＜0.01). Conclusion　 IL-18 may relate to disease activity and the inflammation of liver.

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Study of gastroenteric mucosa immune responses in Balb／c mice by oral immunization with helicobacter pylori

Dongshui LU ; Changqing YU ; Quangming ZOU

Immunological Journal.2001;(2):107-109. doi:10.3969/j.issn.1000-8861.2001.02.009

Objective　To study the mucosa immune responses of gastric and intestinal mucosa in Balb／c mice administered orally with Hp sonicate and mucosal adjuvant（LT）．Methods　The changes of antigen specific AFC in gastric and intestinal mucosa were detected by ELISPOT assay． Results　The numbers of sIgA and IgG AFC rise significantly in PP and gastric mucosa， especially the numbers of sIgA-AFC， significant differences were observed between two immunized groups and the control． Conclusions　Locally synthesized specific sIgA antibodies contribute to immunity against gastric helicobacter infection．

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Construction of mammalian cell expression vector of human CD154 gene from active peripheral blood mononuclear cell and analysis of its sequence

Chunyan ZHANG ; Bo NING ; Shunong LI ; Zhifang ZHANG ; Lianqiang FENG

Immunological Journal.2001;(2):88-90. doi:10.3969/j.issn.1000-8861.2001.02.003

Objective　To obtain mammalian cell expression vector of human CD154 gene. Methods　A 820 bp cDNA fragment was amplified by RT-PCR method from total RNA of human peripheral blood mononuclear cell（PBMC） activated with 10 ng／mL PMA and 1 μg／mL PHA for 8 hours. The fragment was cloned into pcDNA3.1（＋） plasmids.The cloned insert was identified by double digestion of the recombinant plasmid with restriction enzymes BamH Ⅰ and EcoR Ⅰ and sequenced by Sangers-dideory-mediated chain termination. Results　This cDNA fragment included 820 bp entire coding region and a part of the 3 non-coding region. The recombinant mammalian cell expression vector of pcDNA3.1（＋）／hCD154 was constructed， the sequence of the insert was identical to the published sequence encoding human CD154 antigen. Conclusion　The recombinant mammalian cell expression vector of pcDNA3.1（＋）／hCD154 was successfully constructed.

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Development and fabrication of immunosensor

Zhili WEN ; Shiping WANG ; Guoli SHEN ; Xianfang ZENG

Immunological Journal.2001;(2):146-149. doi:10.3969/j.issn.1000-8861.2001.02.022

In this article， several kinds of common immunosensors and the development of their transducers are introduced． Meanwhile， some problems in the fabrication of immunosensor such as immobilization method and reproduction are discussed．

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Display of human decay-accelerating factor on the surface of yeast

Bo GUO ; Peirong XIE ; Qiang ZOU ; Ping ZHEN

Immunological Journal.2001;(2):141-143. doi:10.3969/j.issn.1000-8861.2001.02.020

Objective　To display efficient folding hDAF on the surface of yeast. Methods　The hDAF open reading frame was cloned by PCR from DAF- pBluescript M13-（Amp＋）plasmid， then subcloned into the yeast surface displayed vector pYD1.The recombinant vector was transformed into yeast cells EBY100.Flow cytometric analysis was carried out to evaluate direct binding of anti-DAF mAbs onto the surface of yeast cells displaying DAF. Results　Three mAbs against DAF different epitopes could bind onto DAF displayed on the surface of yeast.Conclusion　Efficient folding DAF can be displayed on the surface of yeast potentially leading to the development of novel applications involving DAF yeast display.

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Study on the correlation of alpha fetoprotein half-life and relapse or metastasis of primary hepatocellular carcinoma

Shulin LIU ; Weimin QI ; Bin LI ; Lijun ZHANG

Immunological Journal.2001;(2):135-137. doi:10.3969/j.issn.1000-8861.2001.02.018

Objective　To clarify the correlation of alpha fetoprotein （AFP） half-life （T1／2） with relapse or／and metastasis of primary hepatocellular carcinoma （PHC） after treatment. Methods　Determination of AFP content in serum with two steps　double McAb sandwich ELISA.Calculation AFP　T1／2 of the sufferers treated with intervention and radiofrequency ablation （RFA） according to T1／2=0.3 dT／log（AFP1／AFP2）. Results　When AFP T1／2＞11.5 days， the sensitivity， specificity and accuracy of predicting the relapse or metastasis of PHC after treatment were 87.5％， 89.3％， 88.5％， respectively. Intervention group was 82.35％， 83.3％， 82.75％， respectively and RFA group was 93.3％， 93.75％， 93.5％， respectively. During 200 days，there　was　very　significant difference of AFP T1／2 （P＜0.001） between non-relapsed／ non-metastasized and relapsed， metastasized and died. Conclusions　AFP T1／2 　can be used as a predictive　index for relapse， metastasis of PHC after treatment. It is not only a reliable parameter for the later stage treatment， but also a useful index for the evaluation of effect.

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