电化学发光免疫分析与酶联免疫吸附试验检测甲型肝炎病毒IgM抗体的对比评价

目的浅议电化学发光免疫分析(ECLIA)与酶联免疫吸附试验(ELISA)检测甲型肝炎病毒(HAV)IgM抗体的对比评价。方法取我院住院或门诊患者血清90例,均用电化学发光免疫分析、酶联免疫吸附试验对甲型肝炎病毒IgM抗体进行测定,并作分析。结果ECLIA、ELISA方式同时对血清标本HAV-IgM抗体在检测时,其阳性符合率为13.33%,阴性符合率=100.00%,总符合率=71.11%。ECLIA敏感度=66.67%,特异度=100.00%,阳性检出率=4.43%。ELISA敏感度=100.00%,特异度=71.43%,阳性检出率=33.33%。结论ECLIA、ELISA对于HAV-IgM抗体的检测中,结果一致性较低,ELISA方式具有较高敏感度,但ECLIA特异性优良,测定时能够全自动化,检验数据更与临床接近,可行实验室测定,为临床疾病诊断供应依据。

酶联免疫吸附试验检测梅毒抗体假阳性的原因分析

分析酶联免疫吸附(ELISA )试验检测梅毒抗体假阳性的原因。方法 本研究抽取一定时间段(2022年1月-2022年12月)期间进行梅毒抗体检测的患者,收集324例病例,回顾性分析资料,以不同的监测方式分为三组,即A、B、C三组,每组108例。其中A组给予ELISA 检测方法,B组采用TPPA检测,C组给予磁微粒化学发光法检测,对三组检测方法进行比较,并分析出现假阳性原因。结果 三组假阳性率中,A组最低,与B、C组相比,差异具有统计学意义,P<0.05;C组假阳性率为29.24%这与B组比较显著要高,差异P<0.05;老年组患者梅毒抗体检测假阳性率计算得出为13.07%显著高于中青年组的6.08%,对比差异明显,P<0.05。结论 梅毒抗体检测采用ELISA方法,年龄、某些疾病、标本污染、标本发生溶血等均是出现假阳性反应的因素,在临床诊疗中,不可单纯依赖实验室结果,需结合患者的具体情况。

Indirect ELISA with Recombinant GP5 for Detecting Antibodies to Porcine Reproductive and Respiratory Syndrome Virus

Porcine reproductive and respiratory syndrome is caused by the PRRS virus (PRRSV),which has six structural proteins (GP2,GP3,GP4,GP5,M and N). GP5 and N protein are important targets for serological detection by enzyme-linked immunosorbent assay (ELISA) and other methods. Toward this goal,we developed an indirect ELISA with recombinant GP5 antigens and this method was validated by comparison to the LSI PRRSV-Ab ELISA kit. The results indicated that the optimal concentration of coated recombinant antigen was 0.2 μg/well for a serum dilution of 1:40. The rate of agreement with the LSI PRRSV-Ab kit was 88.7% (266/300). These results support the potential use of recombinant GP5 as an antigen for indirect ELISA to detect PRRSV antibodies in pigs.

Changes of Nerve Growth Factor in Amniotic Fluid and Correlation with Ventriculomegaly

Objective To detect the change of nerve growth factor (NGF) level in human amniotic fluid during gestation, and to explore the relationship between this change and fetal ventriculo-megaly (VM). Methods The studied subjects (collected from 2004 to 2007) were divided into four groups, including the second-trimester pregnancy group (n=113), third-trimester pregnancy group (n=110), fetal cerebral VM group (n=12), and healthy control group (n=12) which matched with the VM group in gestational weeks. The amniotic fluid specimens were obtained during amniocentesis or cesarean section. The NGF levels in amniotic fluid were detected with en-zyme-linked immunosorbent assay. Results A significantly negative correlation was found between gestational age and the NGF level in amniotic fluid (r= 0.6149, P<0.0001). The NGF level in patients with fetal VM was significantly lower than that in healthy controls (33.95±29.24 pg/mL vs. 64.73±16.21 pg/mL, P=0.024). Conclusion NGF levels in amniotic fluid may be a sensitive marker for fetal VM.

Development of Multiple ELISAs for the Detection of Antibodies against Classical Swine Fever Virus in Pig Sera

The major immunogenic proteins （Ems, E2 and NS3） of classical swine fever virus （CSFV） （Shimen strain） were expressed in E. coli and purified by affinity chromatography. The recombinant antigens were applied to develop multiple enzyme-linked immunosorbent assays （ELISAs） for the detection of specific antibodies in pig sera. Optimum cut-off values were determined by receiver operating characteristic （ROC） analysis after testing 201 sera of vaccinated pigs and 64 negative sera of unvaccinated piglets. The multiple ELISAs were validated with 265 pig sera yielding high sensitivity and specificity in comparison with the virus neutralization results. The results demonstrated that multiple ELISAs can be a valuable tool for the detection of CSFV infection and serological surveys in CSFV-free countries or for the evaluation of the antibody responses in pigs induced by a live attenuated C-strain vaccination

Detection and evaluation of antibodies against neutrophil-activating protein of Helicobacter pylori in patients with gastric cancer

AIM： To detect and evaluate the antibodies against Helicobacter pylori （H pylori） neutrophil-activating protein （HP-NAP） in patients with gastric cancer and other gastroduodenal diseases.METHODS： Recombinant HP-NAP was prepared from a prokaryotic expression system in Escherichia coll. Serum positivity and level of HP-NAP-specific antibodies in sera from 43 patients with gastric cancer, 28 with chronic gastritis, 28 with peptic ulcer, and 89 healthy controls were measured by rHP-NAP-based ELISA. rHP-NAP-stimulated production of interleukin-8 （IL-8） and growth-related oncogene （GROα） cytokines in the culture supernatant of SGC7901 gastric epithelial cells was also detected.RESULTS： The serum positivity and mean absorbancevalue of HP-NAP-specific antibodies in the gastriccancer group （97.7% and 1.01 ± 0.24） were significantly higher than those in the chronic gastritisgroup （85.7% and 0.89 ± 0.14, P 〈 0.005） and healthy control group （27.7% and 0.65 ± 0.18, P 〈 0.001）. The sensitivity and specificity of ELISA for the detection of HP-NAP-specific antibodies were 95.5% and 91.5%, respectively. HP-NAP could slightly upregulate IL-8 production in gastric epithelial cell lines but had no effect on GROα production.CONCLUSION： Infection with virulent H py/ori strains secreting HP-NAP is associated with severe gastroduodenal diseases, and HP-NAP may play a role in the development of gastric carcinoma, rHP-NAP- based ELISA can be used as a new method to detect H pylori infection. The direct effect of HP-NAP on gastric epithelial cells may be limited, but HP-NAP may contribute to inflammatory response or carcinogenesis by activating neutrophils.

Development of a Fast ELISA for the Specific Detection of both Leucomalachite Green and Malachite Green

Malachite green(MG), a dye, is an antifungal agent that has been used to treat and prevent fish diseases. It is metabolized into reduced leucomalachite green forms(LMG) that may reside in fish muscles for a long period, thus being harmful to human health. The aim of this study was to develop a competitive and direct enzyme-linked immunosorbent assay(ELISA) to detect MG and LMG specifically. The monoclonal antibody(m Ab) to LMG was generated using a hybridoma technique. The obtained m Ab showed good cross-reactivity(CR) to malachite green(MG), but not to crystal violet(CV) and Brilliant Green(BG). The m Ab was used to develop a fast detecting ELISA of MG and LMG in fish. By introducing the conjugation LMG-HRP, the detection capability was 0.37 ng m L-1 for MG and LMG. The mean recovery from spiked grass carp tissues ranged from 76.2% to 82.9% and the coefficients of variation varied between 1.8% and 7.5%. The stable and efficient monoclonal cell line obtained is a sustainable source of sensitive and specific antibody to MG and LMG.

Determination of Postvaccinal Antibodies Against OspA B. afzelii, B. garinfi and B. burgdorferi sensu stricto Using Elisa Method

The specific recombinant proteins rOspA B. afzelii, rOspA B. garinii a rOspA B. burgdorferi sensu stricto were used as antigens for construction of ELISA sets. ELISA examination enables determination of specific post-vaccination antibodies against OspA B. afzelii, B. garinii a B. burgdorferi sensu stricto.Using recombinant DNA technology, genes from B. afzelii, B. garinii and B. burgdorferi sensu stricto were inserted into E. coli-expression vectors and the rOspA proteins were produced. These proteins were used for the construction of ELISA kits for the determination of post-vaccination antibodies against individual Borrelia serovars contained in the vaccine. The antibody response of dogs vaccinated with whole-cell vaccine BORRELYM 3 and non-vaccinated dogs was monitored and compared. The ELISA method proved as highly sensitive for the determination of post-vaccination antibodies against individual Borrelia serovars in vaccinated animals. Detection of these antibodies and their quantification may be used for evaluation of efficiency of vaccines against Lyme borreliosis in dogs caused by Borrelia burgdorferi sensu lato. Prospectively, it will be necessary to establish a correlation between post-vaccination antibody levels and protective immunity of vaccinated dogs.