174例乙型肝炎病毒前S1抗原体检测的临床价值分析

目的：对乙型肝炎病毒（HBV）前S1抗原检测的临床价值进行分析.方法：选择在2006年2月至2011年2月来我院就诊的174例乙型肝炎患者作为研究对象,用双抗体的夹心酶联免疫吸附法（ELISA法）检测研究对象血清中的前S1抗原和HBV血清标志物.结果：HBV前S1抗原在HBeAg（乙肝e抗原）阳性组中的检出率高达87.8%,明显高于在HBeAg阴性组中的检出率（32.9%）,根据统计学分析,二者的差异有统计学意义（P＜0.05）.结论：通过对HBV前S1抗原和HBeAg的检测分析,证明了HBV前S1抗原和HBeAg具有非常好的一致性,同时还具有其独特的检测价值,可以补充HBV血清标志物检测中的不足.

抗口腔支原体、精氨酸支原体单克隆抗体的制备及其应用方法的初步研究

用口腔支原体、精氨酸支原体抗原免疫ＢＡＬＧ／ｃ小鼠，取其脾细胞，与小鼠骨髓瘤细胞（Ｓｐ２／０）融合，获得了７株稳定分泌抗口腔支原体单克隆抗体（ＭｃＡｂ），２株分泌抗精氨酸支原体ＭｃＡｂ的杂交瘤细胞株。间接ＥＬＩＳＡ测定其抗体效价可达１：１０３～１：１０７；９株ＭｃＡｂ中１株为ＩｇＧ２ａ，其余均为ＩｇＧ１。初步建立了ＥＬＩＳＡ检测支原体的方法。

食品中常见有害微生物的常规检验方法分析

随着经济的发展,当前人们再食品方面有了更多的选择,但同时食品安全问题也成为了人们关注的重点问题。在生活中,食品从种植、加工到出售,每一个环节都能让食品出现质量问题,尤其是当有害微生物寄存在食品中时,一旦食用含有有害微生物的食品便将会影响食用者的健康甚至生命安全。因此,对食品中常见的有害微生物进行检验成为保证食品安全的重要举措。目前,针对食品中常见有害微生物的检验有多种方法可以检出,不同的检验方法在检验原理、检验方法以及检验结果准确性方面会有一定差异。基于此,本文旨在介绍食品中常见有害微生物的常规检验方法,分析了影响检验结果准确性的因素并提出了提高检验结果准确性的对策。

应用琼脂扩散法检测小鹅瘟抗原抗体

近年来,养鹅业在我省发展十分迅速,但由于鹅种蛋来源复杂,使小鹅瘟在我省呈流行趋势,造成了严重的经济损失。为了更好地防制本病,建立快速、准确、方便的诊断方法十分重要。过去常以病毒分离鉴定和剖检的病理变化进行确诊,需要较长时间,操作也较复杂。另外,检测小鹅瘟免疫...

胃肠道间质瘤的免疫治疗

胃肠间质瘤(GISTs)是最常见胃肠道间叶组织肿瘤,传统放化疗无效,手术完整切除是可能治愈该病的唯一方法。虽然GISTs靶向治疗效果显著,但KIT或PDGFRA基因多位点突变和/或二次突变导致靶向治疗耐药及复发率逐年升高,因此寻找新方法成为治疗GISTs研究热点。GISTs内部存在大量肿瘤浸润免疫细胞和复杂免疫微环境,多种免疫细胞参与GISTs发生、发展,且在靶向治疗中发挥重要作用,GISTs免疫治疗基础及临床试验结果均证实GISTs免疫治疗的可行性及有效性。本文系统总结了GISTs免疫治疗现状及最新研究进展。

Comparative Study of Immunological Properties on Glutamine Synthetase Isozymes in Rice Plants

In rice (Oryza sativa L.) roots two glutamine synthetase (GS) isozymes, GSra and GSrb, were identified recently in the author's experiments, but the homology of both GSra and GSrb as well as their localization in the rice roots are unclear. In the present study, the purified GSra and GSrb from rice roots were used to immunize rabbits to obtain the respective antibodies. The immunodiffusion and immunoblotting experiments showed that the antibody against GSra or GSrb was specific for GS and its isozymes. The immunoprecipitation test indicated that the antibody of GSra or GSrb not only recognized its respective antigen, but also well recognized each other's antigen. GSra or GSrb antibody recognized also better cytosolic GS1 of rice leaves, but the recognization for chloroplast GS2 from rice or spinach (Spinacia oleracea Mill.) leaves was weaker. Our results indicate that GSra and GSrb from rice roots are quite similar in antigenicity and are extremely similar proteins and that both GSra and GSrb may also be a form of cytosolic GS just as the cytosolic GS1 of rice leaves.

Relationship between Vascular Elasticity and Spontaneous Rupture of Hepatocellular Carcinoma

Objective Spontaneous rupture of hepatocellular carcinoma (HCC) is common in Asia and Africa with unclear mechanism. In our previous study, we found that the deposition of immune complex on vascular wall and vascular injury were related to the HCC rupture. In this study, the structure of elastin around the small artery was deeply investigated to confirm our previous study. Methods Immunohistochemical technique and transmission electron microscopy were used to study 23 specimens from ruptured HCC and 30 cases of nonruptured HCC. Results The layer of elastin around the vascular wall was significant thicker in patients with ruptured HCC than that in nonruptured HCC. The proliferation of elastin, abnormal distribution of neutrophil elastase and degradation of collagen fibril were predominantly present in the specimens from ruptured HCC. The phenomenon of electron—dense deposit in the elastic lamina that represented the deposition of immune complex, and the signs of infiltrated neutrophils from bloodstream into the vascular wall that caused the vascular injury, also can be found in ruptured HCC. Since the damaged vessels could become stiff and weak, which would more prone to be splitting and results in hemorrhage and the rupture of HCC, we postulated that the preexisting of immune complex deposition and vascular injury may be relate to the ruptured HCC. Conclusion The vascular injury caused by immune complex deposition might relate to ruptured HCC. Key words hepatocellular carcinoma - rupture - elastin - elastase

Preparation and Identification of Specific Monoclonal Antibody against Porcine Circovirus Type 2

BALB/c mice were immunized using synthetic tandem polypeptide of Cap protein epitope of porcine circovirus type 2 （PCV2） as the antigen. By using lym-phocyte hybridoma technique, a hybridoma cellline stably secreting monoclonal an-tibody against PCV2-rCap protein was successful y obtained and named as 670#. The ascites titer of the obtained monoclonal antibody was 1∶100 000. Western blot results showed that the monoclonal antibody could react with prokaryotical y ex-pressed PET32a-ORF2 recombinant protein, eukaryotical y expressed ORF1-ORF2 tandem protein and PCV2 whole virus celllysate. Indirect EILSA demonstrated that the monoclonal antibody could bind with ORF1-ORF2 tandem protein. Indirect im-munofluorescence assay （IFA） indicated that the monoclonal antibody could identify native PCV2 virus. The preparation of this monoclonal antibody provided technical tools for epitope analysis and molecular diagnosis of PCV2 virus.

In vitro Antimicrobial Activity of Golden Buckwheat Extract and Therapeutic Effect of its Preparation on Chicken Mycoplasma Infection

[Objective] This study aimed to identify the in vitro antibacterial activity of golden buckwheat extract and investigate the therapeutic effect of its preparation on mycoplasma infection. [Method] Through measuring the minimum inhibitory concentra-tion, the in vitro antibacterial activity of golden buckwheat water extract was deter-mined; meanwhile, the therapeutic effect of golden buckwheat oral solution on my-coplasma infection was determined by artificial y infecting chickens with Mycoplasma gal isepticum culture. [Results] The golden buckwheat water extract had obvious in-hibitory effects against Pseudomonas aeruginosa and Escherichia coli, and a certain inhibitory effect on Salmonel a and Staphylococcus aureus; administration of golden buckwheat oral solution at the dose of 0.5%-1.0% continuously for 5 d had a good therapeutic effect against mycoplasma infection. [Conclusion] The study provides sci-entific bases for further study on the antibacterial activity of golden buckwheat and its application.

Generation of Antibodies Against DMRT1 and DMRT4 of Oreochromis aurea and Analysis of Their Expression Profile in Oreochromis aurea Tissues

Sex determination is composed of somatic and germ-line sex differentiation hierarchies whose interaction is poorly understood. A single gene known to control somatic sex determination, the DM-domain containing （Doublesex/Mab-3 DNA-binding motif） gene, is highly conserved across species. Vertebrate DMRT1 （DM-related transcription factor 1） expression occurs predominantly in the testis. Here, however, isolated two distinct DM-domain cDNA from Oreochromis aurea ovary and testis have been named DMRT4 （DM-related transcription factor 4） and DMRT1 by BLAST, respectively. Despite high homology in the DM-domain there is little similarity outside the DM-domain.To better understand the structure, function, and possible roles of DMRT4 and DMRT1 as potential candidates for sex differentiation and sex determination, the intact regions encoding DMRT4 and DMRT1 obtained by PCR were sub-cloned into the vector pMAL-c2x and introduced into the Escherichia coli TB1 cell for efficient fusion expression. After purification and cleavage, DMRT4 and DMRT1 proteins were used to immunize adult rabbits following standard protocols. Consequently, it was found by using Western blot analysis that polyclonal antibodies against DMRT4 and DMRT1 had high specificity. The relative expression levels of DMRT4 and DMRT1 mRNA were determined by fluorescent Real-time RT-PCR in female and male Oreochromis aurea with 13-actin as the internal standard. DMRT1 was expressed only in testis, whereas DMRT4 was over expressed in the ovary, but in both female and male, a slight expression in the brain was also detected. Statistical analysis showed that in the brain, mean DMRT4 mRNA levels in female were significantly higher than in male. Meanwhile, the expression of DMRT4 and DMRT1 protein was also analyzed using the purified antibodies through Western blot and immunohistochemistry. It was found that DMRT4 was exclusively expressed in the ovary and DMRT1 in the testis. Study on DMRT4 and DMRT1 expression facilitated the elucidation of their roles and the understanding of sex differentiation of fish.

Alterations of red blood cell immunoadherence function in hepatitis B patients

AIMS To investigate the alterations of RBC immunoadherence function in patients with various hepatitis B. METHODS RBCC3bRR,RBCICRR and serum CIC levels were measured in 42 patients with acute and chronic hepatitis B at ac- tive and convalescence stages. RESULTS RBCC3bRRs at the active/acute stage of various hepatitis were decreased.They were 13,54%±5,23% in AH, 7.61%±4.12% in AFH,and 16.18%±6.10% in CH, respectively,all of which were lower than those in normal persons (18.12%±3.91% ).At the quiescent/recovery stage of various hepatitis,the RBCC3bRRs were increased significantly.The changes of RBCICRR and serum CIC level were contrary to those of RBCC3bRR. CONCLUSIONS RBC immunoadherence function is decreased in acute and chronic hepatitis.The decrease is in direct proportion to the severity of the diseases.

Identification of Plasma Membrane Aquaporin in Guard Cells of Vicia faba and Its Role in Stomatal Movement

Water channels or aquaporins are the main pathways of water transport. Both the existence and function of aquaporins in die guard cells of Vicia faba L. were investigated both by using RD28 cDNA and RD28 antibody as probes, and by controlling stomatal movement as a parameter combined with antibody and inhibitor of aquaporins respectively. The results revealed that RD28 mRNA, encoding a plasma membrane aquaporin, expressed in ale mesophyll cells and vascular tissues of V. faba, especially in guard cells. And the location of RD28-like proteins was mainly on plasma membrane of guard cells. The addition of 25 mumol/L HgCl2, an aquaporin blocker, and antibody of RD28 as well, greatly suppressed the stomatal opening or guardcell protoplast swelling induced by fusicoccin and light, and closing induced by abscisic acid. However, 5 mmol/L, beta-mercaptoethanol, a reverse reagent of aquaporin blocker, reversed the inhibitory effect of HgCl2 Pretreatment oil stomatal opening ( i.e., HgCl2 was removed after HgCl2 pretreatment for 10 min). The results suggest that the aquaporins in V. faba are associated with stomatal movement.

Antibody Detection on Mycoplasma capricolum subsp. Capripneumoniae in Qinghai Province

[Objective] The aim was to investigate the prevalence of Mycoplasma capricolum subsp. Capripneumoniae in Qinghai Province. [Method] By using indirect hemagglutination test kit for detecting Mycoplasma capricolum subsp. Capripneumoniae,208 goat serums were detected. [Result] The positive rate of goat sera was 16.3%,and the positive rate of sera from different regions ranged from 6.7% to 24.3%. [Conclusion] The infection rate of Mycoplasma capricolum subsp. Capripneumoniae was high in Qinghai Province,so it is necessary to strengthen the prevention and control of this disease.

Study on Drug Resistance and Relative Mechanisms of Chlamydia Trachomatis

Chlamydia Trachomatis (C.T.) is one of the most common pathogens of human sexually transmitted diseases. Treatment of C.T. infection primarily depends on Tetracyclines, Macrolides and Quinolones, but with the wide use of antibiotics an increasing number of drug-resistant Chlamydia trachomatis cases have been reported. This review summarizes the resistant conditions and the possible resistance mechanisms of C.T..

Expression and Purification of E.coli NrfA Protein and Preparation of Polyclonal Antibody against NrfA

[Objective] This study aimed to clone the E. coil NrfA gene and construct the pET-28a （＋）-NrfA prokaryotic expression vector for preparation of polyclonal anti- body against E. coil NrfA. [Method] E. coil NrfA gene was cloned from the E. coli genome DNA by PCR and inserted into the vector pET-28a（＋） to construct prokary- otic expression vector pET-28a （＋）-NrfA. E. coil NrfA protein was expressed by IPTG induction and purified. Polyclonal antibody against NrfA protein was prepared by im- munizing rabbit with routine method. The specificity and titer of polyclonal antibody was confirmed by ELISA and Western Blotting. [Result] The constructed prokaryotic expression vector pET-28a（＋）-NrfA was induced by IPTG, the recombinant NrfA pro- tein could be expressed effectively. The titer of rabbit anti-NrfA polyclonal antibody obtained by immunization and purification was about 1：204 900. Western Blotting anal- ysis indicated that the obtained polyclonal antibody against E. coil NrfA protein had high titer and high specificity. [Conclusion] E. coil NrfA gene was cloned and the prokaryotic expression vector pET-28a （＋）-NrfA was constructed successfully, poly- clonal antibody with high titer and high specificity was prepared, which laid the foun- dation for the study of NrfA in different strains of bacteria.

Analysis of BAC_5 mcAb-Related Epitope Using Random Peptide library

Objective To identify epitope relating to BAC 5 mcAb, a kind of monoclonal antibody (mcAb) located on the surface of nasopharyngeal carcinoma (NPC) cells. Methods Using BAC 5 mcAb as a selected target, the 3 rounds of biopanning to a 12 mer random peptide library (RPL) presented by M13 phages were carried out. The positive M13 phage clones were chosen and confirmed with sandwich ELISA for antibody capture and competitive assay. The exogenous DNA fragments in the positive/negative M13 phages were sequenced to deduce and compare the order of the amino acids of exogenous peptides among the phage clones. Results 77% (35/45) of the phages eluted from the 3rd round of biopnning could be captured by BAC 5 mcAb. The 3 kinds of the peptides were displayed by M13 phages from the 8 positive clones identified with competitive assay. The same character of '-P-V-'structure existed near N-terminus of the 3 different peptides, i.e. -H-Q-S-H-Y-P-Y-P-V-V-S-L- (4/8) -Q-N-Q-A-W-F-S-Q-P-V-R-M- (3/8) and T-Q-A-Y-K-G-F-P-V-L-P-S- (1/8) in comparison with the peptide ' -N-H-Q-S-T-F-W-Q-K-W-T-A-' displayed by M13 phages from the negative clones (6/6). Conclusion BAC 5 mcAb can recognize the 3 kinds of the peptides with-P-V-structure near N-terminus. These peptides mimic the structure of the epitope on the surface of NPC cells recognized by BAC 5 mcAb.

Efficacy of current guidelines for the treatment of spontaneous bacterial peritonitis in the clinical practice

AIM： To verify the validity of the International Ascites Club guidelines for treatment of spontaneous bacterial peritonitis （SBP） in clinical practice. METHODS： All SBP episodes occurring in a group of consecutive cirrhotics were managed accordingly and included in the study. SBP was diagnosed when the ascitic fluid polymorphonuclear （PIN） cell count was 〉 250 cells/mm^3, and empirically treated with cefotaxime. RESULTS： Thirty-eight SBP episodes occurred in 32 cirrhotics （22 men/20 women; mean age： 58.6 ＋ 22.2 years）. Prevalence of SBP, in our population, was 27%. Ascitic fluid culture was positive in nine （24%） cases only. Eleven episodes were nosocomial and 71% community-acquired. Treatment with cefotaxime was successful in 59% of cases, while 41% of episodes required a modification of the initial antibiotic therapy because of a less-than 25% decrease in ascitic PMN count at 48 h. Change of antibiotic therapy led to the resolution of infection in 87% of episodes. Among the cases with positive culture, the initial antibiotic therapy with cefotaxime failed at a percentage （44%） similar to that of the whole series. In these cases, the isolated organisms were either resistant or with an inherent insufficient susceptibility to cefotaxime. CONCLUSION： In clinical practice, ascitic PMN count is a valid tool for starting a prompt antibiotic treatment andevaluating its efficacy. The initial treatment with cefotaxime failed more frequently than expected. An increase in healthcare-related infections with antibiotic-resistant pathogens may explain this finding. A different first-line antibiotic treatment should be investigated.

Diversity of Helicobacter pylori isolates in expression of antigens and induction of antibodies

AIM: To obtain evidence for selection of antigens used in genetically engineered vaccine against Helicobacter pylori (H pylori). METHODS: Enzyme linked immunoabsorbent assay (ELISA) was established on the basis of recombinant protein antigens rUreB, rHpaA, rVacA, rCagA1, rNapA, rFlaA and rFlaB of H pylori to detect expression rates of the antigens in bacterial isolates as well as positive rates of the antibodies in sera from H pylori-infected patients. PCR was applied to the detection of carrying rates of the genes encoding antigens in the isolates. RESULTS: The outputs of rUreB, rHpaA, rVacA, rCagA1, rNapA, rFlaA and rFlaB were approximately 35%, 32%, 15%, 23%, 56%, 25% and 20% of the total bacterial proteins, respectively. One hundred and fifty-one strains of H pylori were isolated from 347 biopsy specimens of chronic gastritis, peptic ulcer or gastric adenocarcinoma, with a positive rate of 43.5%. All of the isolates expressed UreB, HpaA, FlaA and FlaB while 52.3%, 92.1% and 93.4% of the isolates expressed VacA, CagA and NapA, respectively. In the sera of 151 H pylori-infected patients, the positive ratesof IgG antibodies against UreB, HpaA, VacA, CagA, NapA, FlaA and FlaB were 100%, 87.4%, 43%, 71.5%, 89.4%, 84.8% and 79.5%, respectively. Furthermore, the expression frequencies of VacA and NapA were found to be relative to the severity of gastric diseases (P = 0.016 and P < 0.0001, respectively). CONCLUSION: UreB antigen is the top option of developing genetically engineered vaccine against H pylori followed by NapA or HpaA.

Anti-CD132 Monoclonal Antibodies Inducing T Cells Apoptosis after Alloantigen Stimulation and Its Possible Clinical Applications

Aim To investigate the mechanism of anti-CD132 monoclonal antibodies (mAbs)inhibiting T cells proliferation in vitro, and their potential values for clinical use. MethodsBALB/c and C57BL/6 mice splenocytes were harvested for two-ways mixed lymphocyte culture (MLC).Anti-CD132 mAbs (final concentration 100 mg·L^(-1)) were added in MLC on day 0 (group 1) or day 3(group 2). Fluorescence activated cell sorting (FACS) was used to measure the proliferation(carboxy-fluorescein dia cetate, succinimidyl ester, CFSE), apoptosis of T cells (PE-CD3,FTTC-annexin-v), and cell cycle (pro-pidium iodide stain) . The expression of survivin in T cellswas detected by immunochemical stai-ning. Re-sults Multi-generation CFSE-labeled splenocytes werefound dividing and their fluorescent strength decreased in MLC. There was no noticeable change influorescent intensity in group 1 and group 2. On day 3, apoptosis induced by anti-CD132 mAbs wasdetected in part of T cells, but was not detected in the former two days in group 1. In group 2, thenumber of cells in M phase (activated T cells) decreased and apoptot-ic cells increased on day 4.The phenomena were not observed in control group (P < 0.01). Expression of survivin in T cells wasdetected in control group but not in groups 1 and 2. Conclusion Blockade of CD132 signaling pathwayinhibits T cell proliferation in vitro by means of inducing activated alloreactive T cell apoptosisbut not the resting T cells. Anti-CD132 mAbs may be candidates for clinical applications.