HFRS患者汉坦病毒特异性抗体的动态变化

肾综合征出血热（HFRS）在我国危害严重，我们利用汉坦病毒重组核蛋白（rNP）和重组糖蛋白（rGP）为抗原，对25例急性期HFRS患者的90份血清进行了特异性IgA、IgM检测，探讨HFRS患者的抗体动态变化规律，为临床诊断和治疗提供理论依据。     

重组杆状病毒表达汉坦病毒结构蛋白在实验动物中的免疫应答

用杆状病毒载体表达汉坦病毒核壳蛋白(rNP)和糖蛋白(rGP)免疫CBA/J小鼠和金黄地鼠,观察了表达产物刺激产生抗汉坦病毒体液免疫、细胞免疫及免疫保护作用。研究发现,所有免疫小鼠均产生抗汉坦病毒抗体;rNP加rGP共同免疫组小鼠血清IgG抗体滴度明显高于单独rNP或rGP免疫组。同时,共同免疫组中和抗体滴度达1:28或更高,而单独rGP免疫组6只小鼠中只有3只检测到低滴度中和抗体(1:10),rNP免疫组未检测出中和抗体。结果提示,rNP对rGP诱发产生抗病毒中和抗体具有某种协同作用。淋巴细胞转化实验证实,rNP和rGP均能刺激汉坦病毒特异性淋巴细胞增殖反应。以地鼠感染模型对病毒攻击的免疫保护试验表明,rNP和rGP免疫均能使地鼠获得免疫保护。     

肾综合征出血热IgM抗体早期诊断两步法MacELISA的建立

目的 建立和改善肾综合征出血热早期诊断IgM抗体捕获MacELISA法.方法 汉坦病毒核蛋白重组表达纯化后用辣根过氧化物酶标记,建立一种以酶标抗原为基础的MacELISA(称为两步法MacELISA)并与常规三步法MacELISA进行比较分析.结果 检测不同病日HFRS患者血清IgM抗体比较两步法与三步法MacELISA高度相关,敏感性和特异性为100%,无有明显差别.结论 本方法操作简单,用时少,成本低,适合用于肾综合征出血热早期诊断和汉坦病毒感染的监测.     

抗IgG、IgM、IgA型心磷脂抗体和抗β_2糖蛋白1抗体在系统性红斑狼疮患者中的意义

目的探讨抗IgG、IgM、IgA型心磷脂抗体和抗β2糖蛋白1抗体在系统性红斑狼疮(systemic lupus erythematosus,SLE)患者中的意义。方法分别采用化学发光免疫分析法和酶联免疫吸附法检测372例SLE患者、80例其他结缔组织病患者及60例健康体检者血清中抗IgG、IgM、IgA型心磷脂抗体和抗β2糖蛋白1抗体。同时分析这4个指标与SLE及其病情严重程度的相关性。结果在检测的372例SLE患者组中,抗IgG、IgM、IgA型心磷脂抗体和抗β2糖蛋白1抗体的阳性率分别为31.45%、14.52%、10.22%和29.03%,其表达水平分别为(90.39±35.43)U/ml、(41.25±23.16)U/ml、(32.27±15.77)U/ml、(61.42±21.69)U/ml,与SLEDAI积分呈正相关(P0.05)。在80例疾病对照组中,抗IgG、IgM、IgA型心磷脂抗体和抗β2糖蛋白1抗体的阳性率分别为7.50%、2.50%、0和1.25%。而在60例正常对照组中,4项检测结果均为阴性。抗IgG型心磷脂抗体阳性与血小板减少、白细胞减少、血栓形成、习惯性流产和肾脏病变有关;抗IgM型心磷脂抗体阳性与血小板减少、白细胞减少、血栓形成和肾脏病变有关;抗IgA型心磷脂抗体阳性与血小板减少、血栓形成和肾脏病变有关;抗β_2糖蛋白1抗体阳性与血小板减少、白细胞减少、溶血性贫血、血栓形成、习惯性流产和肾脏病变有关。结论抗IgG、IgM、IgA型心磷脂抗体和抗β_2糖蛋白1抗体在SLE的发病及病情发展中起到了非常重要的作用,定量和联合检测对SLE的诊断、病情严重程度的评价、判断预后及疗效观察等均有重要的临床意义。     

化学发光酶联免疫法检测汉坦病毒IgM抗体的研究

目的建立化学发光酶联免疫分析法（CLEIA）检测肾综合征出血热（HFRS）患者血清中IgM抗体。方法以抗人IgM-μ链抗体包被黑色不透明聚乙烯板，辣根过氧化物酶标记汉坦病毒核蛋白作为检测抗原以及luminol-H2O2作为发光底物，建立CLEIA法并对CLEIA与IgM抗体捕获酶联免疫吸附法（MacELISA）进行比较。结果CLEIA与MacELISA相关系数0．97；对于51份确诊的HFRS患者急性期血清CLEIA检测敏感度100％，MacELISA为90．2％；对48份正常人血清两种方法检测特异度均为100％；CLEIA次内变异系数范围5．02％-12．7％，次间变异系数范围0．4％～7．0％，与MacELISA相当。结论化学发光酶联免疫分析法是一种更为灵敏，准确和稳定的方法，适用于检测HFRS早期患者血清中IgM抗体。     

烟台地区肾综合征出血热患者血清抗体检测和汉坦病毒序列测定

目的 了解烟台地区肾综合征出血热(HFRS)患者血清中IgG、IgM抗体水平，确定引起该地区HFRS流行的汉坦病毒的型别分布。方法 收集临床HFRS急性期和恢复期患者血清；用EMSA检测IgG、IgM抗体；用交叉空斑减少中和试验检测中和抗体；采用Trizol法提取患者血清中HFRS病毒RNA，用套式PCR产物做TA试验，测定核苷酸序列。结果 HFRS患者血清IgM阳性率为82．2％(88／107)，IgG阳性率为85．7％(66／77)。该地区两城市38份HFRS患者血清中，有32份属家鼠型病毒(SE0)感染，另6份未能定型；另一城市16份HFRS患者血清中，有15份属姬鼠型病毒感染，1份未定型。该地区HFRS病毒与SE0的同源性达90％以上。结论 引起烟台地区HFRS流行的汉坦病毒，属于以SE0为主的混合型病毒。     

肾综合征出血热患者抗体的消长动态

<正> 肾综合征出血热(HFRS)的早期快速诊断国内已多报道,据认为IgM和IgG类特异性抗体均可作为早期诊断的依据。我们试用2-巯基乙醇(2-ME)处理待检血清,用血凝抑制试验检测IgM,籍以观察HFRS患者不同病期IgM和IgG类抗体的消长动态,结果如下。     

不同亚型磷脂抗体在人群中的诊疗价值

目的 探讨不同亚型(IgG、IgM和IgA型)抗心磷脂抗体(aCL)和不同亚型(IgG、IgM和IgA型)抗β2糖蛋白1抗体(anti-β2GP1抗体)在中国人群中的诊疗价值.方法 采用病例对照研究,选取2019年3月至2020年3月在北京顺义区医院就诊患者共900例(其中抗磷脂综合征45例),40例健康对照者.采用化学发光免疫测定法(CLIA),分别采用IgG、IgM和IgA单亚型试剂检测aCL和aβ2GP1抗体,在不同人群中的分布情况.正态分布数据的结果用均值标准差表示,两组间比较采用t检验,工作特征曲线(ROC)计算曲线下面积(AUC).采用Logistic回归分析确定APS患者aPL同型阳性与临床表现之间的关系.SPSS 24.0对数据进行统计分析.结果 总体aCL患病率为9.23％,其中IgG亚型aCL最常见为8.44％,IgM和IgA亚型aCL患病率仅为2.78％和2.33％.总体aβ2GP1抗体患病率为10.1％,其中IgM亚型aβ2GP1抗体患病率最高为7.56％,其次IgG亚型aβ2GP1抗体患病率为6.11％,最低IgA亚型aβ2GP1抗体患病率为2.11％.IgG、IgM和IgA亚型单独抗体阳性率分别为57.8％、19.2％和3.61％;在aβ2GP1抗体阳性的受试者中,IgM、IgG、IgA单独阳性率分别为51.6％、20.1％和2.19％.与健康对照组相比,APS患者的aPL亚型特异性抗体阳性率明显升高.不同亚型APS抗磷脂抗体结果显示,总体PAPS和SPAPS,aCL和aβ2GP1抗体患病率分别为55.6％和57.8％,与PAPS相比SAPS患者aCL的阳性检出率更高(χ2=16.42,P<0.001;χ2=7.812,P=0.005).结论 IgG、IgM型抗心磷脂抗体和aβ2GP1抗体在诊断及判断病情程度中的应用优于其他亚型,IgA型抗心磷脂抗体和IgA抗β2糖蛋白1抗体为临床的诊疗价值极低,无论在实验室还是在临床实践中,IgA aPL的存在并没有为中国人群的APS诊断提供附加价值.     

细胞因子及特异性抗体的检测在肾综合征出血热诊断中的意义

探讨检测血清细胞因子及肾综合征出血热(HFRS) 病毒特异性抗体IgM和IgG的含量在HFRS发病机制及诊断中的意义.选择24例HFRS患者及30例健康人血清标本,采用生物素-亲和素-酶免疫技术检测IL-2、IL-6和TNF-α,ELISA方法检测血清HFRS病毒特异性抗体IgM和IgG,并对其进行统计学分析. 结果显示, ELISA法检测HFRS患者抗HFRS病毒IgM和IgG的阳性率分别为75.00 % 和50.00 %,健康对照组的抗体阳性率为零;HFRS患者血清IL-2、IL-6、TNF-α的含量分别为10.88±2.31pg/mL、256.46±102.51pg/mL和45.63±5.32pg/mL,高于健康对照组0.59±0.24pg/mL(P＜0.01)、53.8±19.21 pg/mL(P＜0.01)和5.81±3.58 pg/mL(P＜0.01). 结论 :HFRS患者血清IL-2、IL-6和TNF-α及血清特异性抗体IgM和IgG的含量较健康人明显升高,检测这些指标对该病发病机理、诊断及预后评价有一定意义.     

系统性红斑狼疮患者抗β2-糖蛋白Ⅰ抗体检测及其临床意义

目的 探求抗β2-糖蛋白Ⅰ(β2glyeoprotein Ⅰ,β2-GP Ⅰ抗体IgG、IgA、IgM在系统性红斑狼疮(SLE)临床的测定.方法 采用酶联免疫吸附试验(ELISA)检测100例SLE患者、39例疾病对照组(包括类风湿关节炎、硬化症、干燥综合征、自身免疫性肝病和混合性结缔组织病)和30例正常人群血清抗β2-GP 1抗体、抗心磷脂抗体(ACL)IgC、IgA、IgM等指标,分析其与临床特点(如血栓、流产)的关系.结果 SLE组抗β2-GP Ⅰ抗体(IgG、IgA、IgM)浓度高于正常对照组,差异具有统计学意义(P<0.01),敏感性、特异性、阳性预测值、阴性预测值分别为17.2%、95.7%、85.0%、44.6%.β2-GPⅠ抗体(IgG、IgA、IgM)与ACL抗体(IgG、IgA、IgM)两者呈正相关(r值分别为0.418、0.624、0.518、0.583,P<0.01).以SLE DAI为因变量对β2-GPⅠ抗体、ACL抗体、dsDNA、u1-RNP、Sm、SSA、SSB、Sc1-70、Jo-1、P.蛋白、PT、APTT进行Logistic回归统计分析,入选的自变量有β2-GPⅠ·IgC和dsDNA.结论 抗β2GP Ⅰ抗体在SLE中具有一定敏感性和较高的特异性,与SLE的血栓形成相关,且抗β2-CPⅠ抗体IgG是SLE疾病活动性危险因素之一.检测抗β2-GPⅠ抗体在SLE中存在一定临床应用价值.     

Class-specific antibody determination against Aspergillus fumigatus by means of the enzyme-linked immunosorbent assay. III. Comparative study: IgG, IgA, IgM ELISA titers, precipitating antibodies and IgE binding after fractionation of the antigen

IgG, IgA and IgM ELISA antibody titers against Aspergillus fumigatus were elevated in sera of patients with aspergilloma and allergic bronchopulmonary aspergillosis (ABPA), showing higher titers for the IgG antibodies compared with the IgA and IgM antibodies. No differences were found between titers of identical antibody classes in the two groups of sera. IgG and IgA ELISA titers were highly specific whereas IgM ELISA showed more unspecific binding of IgM antibodies. Antibodies, as measured by ELISA, studied after fractionation of the antigen into fractions of decreasing molecular weight, showed a preferential binding by the high molecular weight fractions. Precipitating antibodies studied in patient sera did not always correspond with the IgG ELISA titers. IgE antibody binding was observed in all fractions from Sephadex G-100 fractionated components; maximum binding was found with fractions of 28,000-60,000 daltons. The low molecular weight fractions (18,000-less than 5,000 daltons) showed less IgE binding but the quantity of this fraction was higher. The discrepancies noted between the IgG and IgA ELISA titers and the binding of IgM or IgE antibodies indicate that antigenic components may in part differ in the binding of antibody classes.     

Development of serum and intestinal antibody response to rotavirus after naturally acquired rotavirus infection in man

The temporal characteristics of the response of rotavirus specific IgM, IgG, IgA in serum and secretory antibody in feces to rotavirus were studied in 77 hospitalized patients with rotavirus induced gastroenteritis. The response in serum was characterized by the sequential appearance of rotavirus specific IgM, IgG, and IgA antibody. The IgM antibody appeared to be higher in the acute phase of the disease and was subsequently replaced by the IgG and IgA antibodies. However, the titers of IgG rotavirus antibody in convalescent specimens of serum were found to be statistically significantly lower in patients with severe or prolonged rotavirus infection than in specimens from subjects with mild or moderate disease. Most fecal specimens collected during both the acute and convalescent phase of illness contained virus specific secretory IgA. Higher concentrations of antibody were measured in convalescent samples from patients with prolonged diarrhea and virus shedding. These observations suggest a possible relationship between the severity of rotavirus infection and the nature of systemic and secretory antibody response.     

Campylobacter etiology in human gastroenteritis demonstrated by antibodies to acid extract antigen

Campylobacter antibodies of the immunoglobulin G (IgG), IgM, and IgA classes were determined by enzyme immunoassay with acid glycine extract antigen in patients and controls involved in two Campylobacter outbreaks and in 266 unselected patients with acute enteritis. The assay showed a specificity of 99% for each immunoglobulin class in sera from 200 healthy blood donors. Elevated Campylobacter antibody titers were shown in 97% of stool culture-positive patients involved in the outbreaks. Rapid changes of IgA and IgM Campylobacter antibodies were typical of the early phase of serologic response in the outbreaks and thus offered the best diagnostic value in the serologic diagnosis of acute campylobacteriosis. In unselected patients with acute enteritis, the assay revealed elevated Campylobacter antibody titers in 37 patients, of whom only 12 had had positive Campylobacter stool cultures. In the sera of patients with other bacterial findings in addition to high titers of Campylobacter antibodies, no cross-reacting antibodies were found, but there was evidence of several mixed infections.     

Measurement of Chlamydia pneumoniae-specific immunoglobulin A (IgA) antibodies by the microimmunofluorescence (MIF) method: comparison of seven fluorescein-labeled anti-human IgA conjugates in an in-house MIF test using one commercial MIF and one enzyme immunoassay kit

For the serological diagnosis of acute Chlamydia pneumoniae infection, the microimmunofluorescence (MIF) test is the most commonly used method and also the "gold standard" for the measurement of immunoglobulin G (IgG) and IgM antibodies. The role of IgA antibodies in diagnosis has not been established. Commercially available fluorescein-labeled anti-human IgA conjugates have not been systematically compared to each other, and this situation may cause considerable variations in IgA results. Therefore, we tested 261 serum samples from 122 patients with pneumonia for IgA antibodies by using six alpha-chain-specific anti-IgA conjugates in our in-house MIF test, one commercial MIF test, and one enzyme immunoassay (EIA). Interfering IgG antibodies were removed with Gullsorb reagent before the measurement of IgA antibodies. Altogether, 14 significant IgA antibody increases in serum samples between the acute phase and the convalescent phase were detected by at least one of the conjugates in the MIF test, while no increases were found in the IgA EIA. Only one patient showed a significant IgA antibody increase with all of the fluorescein-labeled conjugates. Five significant titer changes were detected by at least two conjugates, and in nine instances, the titer increase was detected by one conjugate only. The titer agreement indicated by kappa coefficients was very good or good for all of the fluorescein-labeled conjugates and the EIA with low antibody titers but decreased with increasing titers.     

Human serum antibody response inCampylobacter jejuni enteritis as measured by enzyme-linked immunosorbent assay

An ELISA for detection of IgG, IgA, and IgM antibody using an acid-glycine extract from Campylobacter jejuni as antigen was developed. To determine the value of this assay for the diagnosis of acute Campylobacter jejuni infections, the IgG, IgA, and IgM immune response against Campylobacter jejuni was investigated at various timepoints after infection in patients with culture-proven infection. A total of 112 sera from 46 patients and 78 sera from a control group were tested. All but one of the 46 patients with culture-proven Campylobacter jejuni enteritis developed IgG antibodies against Campylobacter jejuni. IgA and IgM ELISA both showed 97% specificity, and sensitivity of 63% and 30% respectively. IgG antibody titers generally remained at a constant level for more than 50 days, whereas IgA and IgM antibody titers declined more rapidly to normal values within 30 to 50 days after onset of clinical symptoms. Detection of Campylobacter jejuni specific IgA antibodies in a single serum sample provided the most useful assay for serological diagnosis of Campylobacter jejuni enteritis. The presence of Campylobacter jejuni specific IgM antibodies was the sole diagnostic criterion in three cases. Serological diagnosis of Campylobacter jejuni enteritis should therefore include both IgA and IgM antibody determination.     

Serum antibody response to filamentous hemagglutinin in patients with clinical pertussis measured by an enzyme-linked immunosorbent assay

Titers of antibodies to filamentous hemagglutinin (FHA) were determined by enzymelinked immunosorbent assay in acute and convalescent phase serum samples from 158 patients with clinical symptoms typical of whooping-cough. In 96 of the patients the diagnosis was verified by culture. Significant changes in serum levels of IgG, IgM and/or IgA antibodies against FHA were demonstrated in 126 patients (80%). Thus, demonstration of significant changes in FHA antibody titers in serum can be used for serological diagnosis of pertussis. The results also show that high levels of IgG, IgM and/or IgA antibodies in a single serum sample suggest current pertussis infection, but if the diagnosis is based on determinations of FHA antibody titers in a single serum sample the sensitivity is low. The levels of antibody to FHA were compared with previously determined levels of antibodies to pertussis toxin. A significant antibody response against both FHA and pertussis toxin was seen in 111 patients (70 %) while 147 patients (93 %) developed a significant increase in antibodies against one or both antigens.     

Measurement of Chlamydia pneumoniae-Specific Immunoglobulin A (IgA) Antibodies by the Microimmunofluorescence (MIF) Method: Comparison of Seven Fluorescein-Labeled Anti-Human IgA Conjugates in an In-House MIF Test Using One Commercial MIF and One Enzyme Immunoassay Kit

For the serological diagnosis of acute Chlamydia pneumoniae infection, the microimmunofluorescence (MIF) test is the most commonly used method and also the “gold standard” for the measurement of immunoglobulin G (IgG) and IgM antibodies. The role of IgA antibodies in diagnosis has not been established. Commercially available fluorescein-labeled anti-human IgA conjugates have not been systematically compared to each other, and this situation may cause considerable variations in IgA results. Therefore, we tested 261 serum samples from 122 patients with pneumonia for IgA antibodies by using six α-chain-specific anti-IgA conjugates in our in-house MIF test, one commercial MIF test, and one enzyme immunoassay (EIA). Interfering IgG antibodies were removed with Gullsorb reagent before the measurement of IgA antibodies. Altogether, 14 significant IgA antibody increases in serum samples between the acute phase and the convalescent phase were detected by at least one of the conjugates in the MIF test, while no increases were found in the IgA EIA. Only one patient showed a significant IgA antibody increase with all of the fluorescein-labeled conjugates. Five significant titer changes were detected by at least two conjugates, and in nine instances, the titer increase was detected by one conjugate only. The titer agreement indicated by kappa coefficients was very good or good for all of the fluorescein-labeled conjugates and the EIA with low antibody titers but decreased with increasing titers.     

Evaluation of antibodies reactive with Campylobacter jejuni in Egyptian diarrhea patients.

Serum and stool samples were collected from 128 individuals: 96 diarrhea patients and 32 apparently healthy controls. Stool specimens were cultured for enteric bacterial pathogens, while sera were screened by enzyme-linked immunosorbent assay for Campylobacter jejuni-reactive antibodies. Of 28 diarrhea patients who demonstrated C. jejuni-reactive antibodies (titers, > 100), 14 were culture positive for this organism. The 32 healthy controls showed significantly lower antibody titers (P < 0.05) with the exception of 10 subjects who were culture positive for C. jejuni and had reactive immunoglobulin M (IgM) (6 subjects) and IgG (7 subjects). IgA was not detected in those 10 individuals (asymptomatic). Avidity was expressed as the thiocyanate ion concentration required to inhibit 50% of the bound antibodies. The avidity was higher in symptomatic patients than asymptomatic healthy controls. IgG was less avid (0.92 M) compared to IgM (0.1 M) and IgA (1.1 M), with no correlation between antibody titer and avidity. However, the thiocyanate ion concentration required for the complete inhibition of IgG (5 M)-bound antibodies was higher than that of IgA (2 M) and IgM (3 M). This study also shows that C. jejuni antibodies were variably cross-reactive with Escherichia coli, Shigella flexneri, Shigella sonnei, and Neisseria meningitidis in addition to Campylobacter coli and Campylobacter rectus.     

Aberrant synthesis of antibodies directed at the Fab fragment of IgA in patients with IgA nephropathies

The sera of 37 patients with IgA nephropathy (IgA NP) were assayed for levels of antibodies specific for the Fab fragment of homologous IgA, and the values obtained were compared to antibody levels in a panel of 26 normal volunteers. IgG antibody levels in IgA NP patients were significantly elevated over those of the controls (P less than 0.01); at the same time IgM anti-Fab alpha levels were significantly decreased when compared to the control panel (P less than 0.01). There was no correlation of antibody levels of either isotype with levels of circulating immune complexes; however, IgM antibody levels of IgA NP patients showed a significant negative correlation with severity of renal insufficiency.