改良抗μ链捕捉ELISA法快速诊断汉坦病毒出血热

作者研制了一种检测汉坦病毒(Hanta Vir-us,HV)所致出血热的抗μ链捕捉ELISA 法。该法是用抗人μ链血清IgG 包被微孔板来捕捉出血热病人血清的抗出血热病毒特异性IgM 后,再加重组的汉坦病毒(HV)核衣壳抗原或Puumula 血清型株(H(?)lln(?)s B1)病毒核衣壳抗原,洗后再加用POD标记的单克隆抗体(抗HV 或抗B1病毒)、整个EL-ISA 试验只需时4小时。     

青岛市流行性出血热的汉坦病毒检测和其基因型的研究

目的探讨引起青岛市流行性出血热的汉坦病毒(Hantavirus,HV)的感染类型,以明确基因亚型和进化特点。方法免疫胶体金法检测126份临床怀疑急性期病人血清中汉坦病毒的抗体。RT-PCR扩增病毒的S基因段,并经测序获得基因序列。病毒的核苷酸序列分析和构建系统进化树应用DNASta软件,并以确定病毒的型别。结果 126份血清标本中,112份汉坦病毒IgM和(或)IgG抗体阳性。经RT-PCR扩增获得7株汉坦病毒S基因序列,与汉滩型(Hantaan virus,HTN)和汉城型(Seoul virus,SEO)毒株比较,其中2株为HTN型,5株为SEO型,相互间序列达到68%-98.9%同源性。结论引起青岛市流行性出血热的汉坦病毒为汉滩型和汉城型的混合感染,以汉城型为主。     

一起流行性出血热疫情的病原学检测与分析

目的对2014年湖北省咸宁市嘉鱼县一起流行性出血热(EHF)疫情的病例血清标本及患者居住地周围捕获的动物鼠的鼠血、鼠肺组织开展汉坦病毒病原学检测及病毒的分子进化研究。方法采用汉坦病毒IgG和IgM抗体检测试剂对2例患者及56例密切接触者开展血清学和核酸检测。对于核酸阳性的标本进一步分型确定其基因型别。在此基础上,对M片段基因序列进行测序并构建进化树。结果 2份送检的患者血清IgM抗体均为阳性,所有密切接触者血清IgM和IgG均为阴性,汉坦病毒核酸全部为阴性;在捕获的8只鼠标本中,3只鼠检出汉坦病毒核酸,阳性率为37.5%,基因分型结果为汉城型(SEO)型。M片段测序及系统发生情况表明该地鼠携带的汉坦病毒位于汉城病毒(SEOV)所在的支系,进化上同武汉分离株WuhanRf02、江西分离株JiangxiXinjianRn-07-2011D亲缘关系最近。结论通过对病例标本及居住地动物鼠进行病原学检测与分析,发现该次出血热疫情很有可能是由SEO型汉坦病毒经鼠造成向人传播。因此,通过加强对该地区汉坦病毒宿主动物的监测对于EHF的防控十分重要。     

IgG2和IgA共同缺乏的献血员

本文作者对44份来自美国6个血液中心的已知IgA缺乏的献血者血清和44份含有抗-IgA的IgA缺乏的病人血清采用特异性的放射免疫扩散法检测了IgG2的含量，并随机抽取20份献血员血清作为对照，且进行了有关的随访。结果显示，44份IgA缺乏的献血员血清中有4份（9．0％）被检出了IgG2缺乏．IgG2含量分别为＜8．0．＜8.0，23．0．28．0mg／dl（正常值为110～800ms／dl）。44价病人血清有14份（31．5％）被检出了IgG2缺乏．是IgA缺乏的献血员的3．5倍（P＜0．01）。对照组20份均在正常范围。随访4例IgG2和IgA共同缺乏献血员．发现3例有反复…     

江门市传染性非典型肺炎(SARS)的血清流行病学调查研究

目的分析江门市各类人群SARS冠状病毒血清IgG抗体水平。方法采集不同人群血标本,检测血清SARS冠状病毒IgG抗体。结果检测各类人群血清标本208份,SARS冠状病毒血清IgG抗体阳性53份,阳性率为25.48%;其中临床确诊SARS病人和高暴露人群阳性率分别为100.00%(47/47)和11.54%(6/52);临床发热的疑似病人及健康人群血标本,检测血清SARS冠状病毒IgG抗体,结果均为阴性。结论SARS冠状病毒是江门市本次SARS流行的主要病原体,在这次疫情暴发之前,人群对SARS冠状病毒普遍易感,江门市SARS监测和防控工作任务相当艰巨,面临极大的挑战。     

间接免疫荧光试验在人群埃可30型肠道病毒IgG抗体检测中的应用

目的建立埃可30型肠道病毒(Echovirus30,Echo30)国内流行株血清特异性IgG抗体检测的间接免疫荧光法(IFA),了解人群既往感染情况。方法采用由爆发人群分离得到的病毒株感染人胚肺二倍体细胞(MRC-5)制备抗原片,探讨IFA检测血清Echo30 IgG抗体的方法,并初步用于检测苏北病毒性脑膜炎爆发后收集的该地区儿童血清中Echo30 IgG抗体。结果建立了Echo30肠道病毒血清特异性IgG抗体的IFA检测方法。在44份健康儿童血清中筛检到28份(63.6%)Echo30 IgG抗体阳性,3份患者血清Echo30 IgG抗体均为阳性。结论Echo30国内流行株IgG抗体的IFA操作简便、经济,可用于血清流行病学调查。爆发地区儿童中存在大量的无症状感染者。     

间接免疫荧光试验在人群埃可30型肠道病毒IgG抗体检测中的应用

目的建立埃可30型肠道病毒（Echovirus30，Ech030）国内流行株血清特异性IgG抗体检测的间接免疫荧光法（IFA），了解人群既往感染情况。方法采用由爆发人群分离得到的病毒株感染人胚肺二倍体细胞（MRC-5）制备抗原片，探讨IFA检测血清Ech030 IgG抗体的方法，并初步用于检测苏北病毒性脑膜炎爆发后收集的该地区儿童血清中Ech030 IgG抗体。结果建立了Ech030肠道病毒血清特异性IgG抗体的IFA检测方法。在44份健康儿童血清中筛检到28份（63．6％）Ech030IgG抗体阳性，3份患者血清Ech030IgG抗体均为阳性。结论Ech030国内流行株IgG抗体的IFA操作简便、经济，可用于血清流行病学调查。爆发地区儿童中存在大量的无症状感染者。     

双单链抗体夹心法诊断汉坦病毒感染的初步研究

目的制备检测汉坦病毒感染的双单链抗体夹心ELISA诊断试剂,并予以评价。方法以抗汉坦病毒NP抗原单链抗体包被反应板,以辣根过氧化物酶(HRP)标记大肠埃希菌表达的抗NP抗原单链抗体,并制成双单链抗体夹心诊断试剂,检测56份早期肾综合征出血热(HFRS)患者血清及66份非HFRS对照血清。结果56份HFRS患者血清中,检测出29例阳性,阳性率为51.8%,对照组66例,检测结果均为阴性。结论研制的试剂特异性好,价格低廉。     

性病门诊患者单纯疱疹病毒（herpes simplex virus，HSV）感染状况分析

目的了解性病门诊患者单纯疱疹病毒Ⅰ、Ⅱ型的血清学流行情况。方法对2010年8月到2011年11月在佛山市第一人民医院皮肤科就诊,拟诊为HSV感染患者的血清标本且均同时检测HSV-1和HSV-2两种血清型,共300份血清标本,其中男217份,女83份。用ELISA方法进行检测,由有经验者按照操作说明书进行测定,用X^2分析进行统计学处理。结果300例HSV感染患者中,其平均年龄为32．95岁,其中男性为217例,女性为83例。HSV-1的感染率明显高于HSV-2的,且存在同时感染的情况。结论HSV感染患者多为中青年,且男性明显多于女性,HSV-1感染率高于HSV-2的,且存在二者同时感染,IgG的阳性率明显高于IgM的,所以对于症状不明显的患者,临床医生应同时检测两种血清型且检测抗体时应同时检测2种抗体,以免漏诊。     

献血员IgG2和IgA共同缺乏

本文作者对美国6个血液中心的44份已知IgA缺乏的献血者血清和44份含有抗-IgA的IgA缺乏的病人血清,采用特异性的放射免疫扩散法检测了IgG2的含量,并随机抽取20份献血员血清作为对照,且进行了有关的随访。     

重组汉坦病毒核壳蛋白抗原诊断肾综合征出血热

目的 证实重组汉坦病毒核壳蛋白（ｒＮＰ）对肾综合征出血热的诊断价值。方法 用大肠埃杀菌表达汉坦病毒７６１１８株Ｓ基因获得ｒＮＰ,从该病毒感染细胞裂解液获得天然ＮＰ。在相同的捕捉法酶联免疫吸附测定（ＥＬＩＳＡ）系统中,比较研究ｒＮＰ与天然ＮＰ的抗原功能。用ｒＮＰ抗原的捕捉法ＥＬＩＳＡ检测临床血清,与间接免疫荧光（ＩＦＡ）检测特异性ＩｇＧ的结果相比较。结果 用ｒＮＰ和天然法ＥＬＩＳＡ检测临床血清,与间     

重组汉坦病毒核蛋白抗原用于免疫滴金技术检测肾综合征出血热抗体的研究

目的　探索应用重组汉坦病毒核蛋白 (rNP)的免疫滴金法 (CGIDA)对肾综合征出血热 (hemorrhagicfeverrenalsyndrome ,HFRS)的诊断价值。 方法　制备纯化汉坦病毒核蛋白抗原 ,构建了HTN型汉坦病毒的核心区域 (334个碱基 ) ,克隆至原核表达载体 pGEX 4T 1中进行原核表达及纯化。在相同的CGIDA系统中 ,比较研究rNP与天然汉坦病毒核蛋白 (NP)的抗原功能。并与酶联免疫吸附试验 (ELISA)和间接免疫荧光法 (IFA)对比检测。结果　用rNP和天然NP平行检测HFRSIgM符合率达 89.7% ,HFRSIgG符合率达92 .3% ;CGIDA法检测HFRSIgM的敏感性为 75 .0 % ,特异性为 10 0 % ;CGIDA法检测HFRSIgG的敏感性为 83.1% ,特异性为10 0 %。结论　重组核蛋白的CGIDA对HFRS的早期诊断具有较好的应用价值。     

Shigella flexneri O-antigen specific enzyme immunoassay: a prospective study of class-specific antibody titres against lipopolysaccharide antigens in Vietnamese children and adults with serotype 1b or 2a dysentery

A total of 278 sera were collected from 97 patients with a bacteriologically verified Shigella flexneri serotype 1b or 2a infection. Of the patients, 65 were children below the age of 5 years and admitted to the Department of Infectious Diseases at St. Paul's Hospital, Hanoi, Vietnam; 32 were adults, aged 20–24 years, and infected during a dysentery outbreak at Son Tay technical school, Hanoi. The sera were analysed for their specific immunoglobulin A (IgA), M (IgM) and G (IgG) titres against phenol-water-extracted and chemically defined lipopolysaccharide (LPS) antigens of S. flexneri and other shigellae by an enzyme immunoassay (EIA). The titres estimated in sera from patients were compared with titres seen in sera from age-matched healthy people living in the Tu Liem district, Hanoi. Positive titres were defined as greater than the mean titre+ 2 SD in sera from healthy persons. Children 1–2 years old and infected with S. flexneri serotype 1b responded with significantly elevated IgA titres up to 60 days after falling ill (P-values 0·06 to <0·001). The IgG titres against the homologous S. flexneri serotype 1b were significantly elevated in samples collected up to 6 months after children up to 5 years old had fallen ill (P-values ranging from 0·007 to <0·001). IgM titres were elevated, but not significantly higher than those seen in healthy individuals. The results suggested that children younger than 3 years who responded with IgA, IgG and, to a lesser extent, IgM increases were experiencing their first S. flexneri dysentery, whereas older children and adults who only showed IgG increases had had experience with previous S. flexneri infections and displayed an anamnestic response. In adults, significant titre increases were seen only in individuals with low pre-infection titres. The clinical data and laboratory analyses of the strains suggest that a virulent S. flexneri serotype 1b clone was prevalent in the Hanoi area in 1982–1983. Young children infected with S. flexneri serotype 2a also responded with elevated IgA and IgG titres, but there were too few children to permit statistical analyses.     

Determination by enzyme-linked immunosorbent assay (ELISA) of specific IgG antibody activities for diagnosis of farmer's lung disease

The specific IgG antibodies to thermophilic actinomycetes in the sera from patients with farmer's lung disease were quantitated by an ELISA and the results thereof were compared with those obtained by the standard double immunodiffusion assay (Ouchterlony's method). All sera from patients with farmer's lung disease were precipitin positive to Thermoactinomyces vulgaris and/or Micropolyspora faeni by the Ouchterlony's method, and revealed significantly high levels of IgG antibody activities against these antigens by ELISA. We have found 18 precipitin-positive but asymptomatic subjects in a survey of the dairy farming population of the northern area of IWATE Prefecture. Their sera were precipitin positive to antigens related to the farmer's lung disease, however IgG antibody activities as determined by ELISA to these antigens was significantly lower in the group of asymptomatics than in the group of symptomatics. Precipitin negative dairy farming controls and normal controls living in the urban area revealed low antibody activities as determined by ELISA. From these studies, we concluded that the assay of specific IgG antibody activity to thermophilic actinomyces by ELISA is useful for the diagnosis and screening of farmer's lung disease.     

Improved diagnosis of pulmonary tuberculosis by detection of antibodies against multiple Mycobacterium tuberculosis antigens

Two secreted antigens (38 and 30 kDa) and 1 cytosolic antigen (16 kDa) were purified in our laboratory from Mycobacterium tuberculosis culture filtrate and cytosol using chromatographic/electrophoretic methods. One recombinant antigen (27 kDa, MPT51) expressed in Escherichia coli was also isolated. All the 4 antigens were tested individually for detection of serum IgG, IgA, and IgM (a total of 476 sera from 5 groups) by indirect enzyme-linked immunosorbent assay. Keeping the well-reported 38 kDa as the main candidate, the usefulness of the other antigens, which may add to the test positivity in cases not diagnosed by 38 kDa, was analyzed. The individual antigens ranged in their sensitivity from 57% to 67% (IgG). Addition of other antigen results, with that of 38 kDa, offered a sensitivity of 91% in smear- and culture-positive tuberculosis (TB), 78% in smear-negative culture-confirmed TB, and 97% specificity in normal healthy subjects. IgG antibody to multiple antigens (38, 30, and 16 kDa) may be a sensitive, specific, rapid, and cost-effective test to rule-in clinical suspicion of pulmonary TB.     

High concentrations of serum IgG can cause pitfalls in antiglobulin tests

High levels of serum IgG can increase the reactivity of sera in antiglobulin tests. This phenomenon implies possible false positive reactions in the immunofluorescence test (IF) and in enzyme-linked immunosorbent assays (ELISA). This was demonstrated in model experiments when commercial gammaglobulin in different concentrations of sera with different levels of IgG were tested against unrelated or specific antigens. The reactivity seems to be due to unspecific binding of the Fc part of the IgG molecule. Specificity problems associated with high concentrations of serum IgG are particularly important when analysing sera from tropical countries, where several infections are associated with pronounced hypergammaglobulinaemia.     

Isoelectric focusing of immunoglobulins as a new method of immune response analysis in staphylococcal infections

The study presents a new way of analysis of IgG response to staphylococcal antigens. The method is based on the isoelectrofocusing of human immunoglobulins and after blotting, their reactivity with purified staphylococcal antigens: alpha-toxin, lipase and teichoic acid. The method analyses not only total IgG but also the ‘monoclonal’ levels of IgG subclasses (clones based on the isoelectric points of immunoglobulins). When the pattern of IgG response to particular antigens were compared, a great diversity between patients' sera samples was observed. The qualitative and quantitative assessment of sera IgG fractions differentiated by pH gradient revealed the individual character for each patient. No correlation could be observed between IgG pattern and the type of staphylococcal infection. Analysing the subclass of IgG showed that for protein antigens (alpha-toxin, lipase) it was mainly IgG1 but for carbohydrate antigens (teichoic acid) it was IgG2. No traces of IgG3 and IgG4 fractions were observed.     

Antibiodies of the IgG, IgM, and IgA classes in newborn and adult sera reactive with gram-negative bacteria

Umbilical cord serum and adult serum antibodies reactive with heat-stable somatic antigens of Gram-negative bacteria (Neisseria gonorrhoeae, Escherichia coli, and Salmonella typhosa) were assayed by using an indirect fluorescent antibody test. Reactive IgG, IgM, and IgA antibodies were identified by using fluoresceinconjugated antisera specific for these immunoglobulin classes.IgG antibody titers in cord serum approximated those found in the corresponding maternal sera. IgM and IgA antibodies were present in adult sera but were not demonstrable or were present only in small amounts in cord sera. The presence of IgG and IgM antibodies reactive with Gram-negative bacteria was confirmed by the testing of purified 7S and 19S fractions. In addition, both IgG and IgM reactivities were inhibited by the prior incubation of serum with purified specific lipopolysaccharide preparations.The ubiquity and magnitude of these natural IgG antibodies in the sera of both adults and neonates have apparently eluded detection in previous studies. The use of bactericidal and agglutination tests, which are apparently more sensitive to the presence of IgM than to IgG antibodies, may account for the failure of previous studies to detect adult and cord IgG antibodies reactive with somatic antigens of Gram-negative bacteria. The presence of these IgG antibodies may be correlated with the resistance to infection demonstrated by most newborns as they are challenged by the septic extrauterine environment.     

Serological typing of Pseudomonas aeruginosa--comparison of various antigenic schema.

Serotyping sera for both the agglutination (using IMSUT sera) and slide agglutination (using TIBS sera) were succesfully prepared using Homma's serotype strains. They were proved to be type specific and distinct. The two typing sera (IMSUT and TIBS) were used to determine the correspondence of major O-antigens among the different kinds of serotype schema which are used at present throughout the world. The serotype sera of Lanyi, Liu and Meitert were also used for the same purpose. The relationships among serotypes of the seven schemata were clarified as shown in tables in the text. Cross-reactions were found among several serotypes in each of several serotype schemata was suggested.     

Characterization of autoantibody activities in sera anti-DNA antibody and circulating immune complexes from 12 systemic lupus erythematosus patients

To examine autoantibodies present in patients with active systemic lupus erythematosus (SLE), sera, circulating immune complexes (CIC), and antibodies purified on DNA-immunoadsorbent were tested by enzyme immunoassay. A panel of self-antigens, including DNA, histones (HIS), glomerular basal membrane (GBM), thymus cell extract (TCE), actin (ACT), myosin (MS), and tubulin (TUB), was used to define their specificities. IgM antibodies against all antigens of the panel were detected in sera, CIC, and in antibodies eluted from the DNA-immunoadsorbent and demonstrated a large polyreactivity. IgG antibodies showed restricted activities against DNA, HIS, GBM, and TCE in sera and a large polyreactivity in CIC. Inhibition experiments were performed to assess their mono- or polyreactivities. Among the IgG autoantibody population recognizing DNA, two populations of IgG antibodies were detected in the sera and in the affinity purified anti-DNA: one recognizes DNA, HIS, and GBM, and the other binds to DNA and to cytoskeletal proteins. These autoantibody populations were found in CIC, which also often contained high amounts of IgG antibodies recognizing ACT and MS. A third population of IgG antibody that recognizes only TCE and could not be inhibited by DNA or other antigens was found in serum and CIC. Our data demonstrate the existence of several populations of autoantibody in serum and CIC of SLE patients: (1) IgM polyreactive autoantibodies, (2) IgG polyreactive autoantibodies recognizing DNA and cytoskeletal proteins, (3) IgG specific to DNA, which cross react with HIS and GBM, and (4) IgG specific to TCE antigens. © 1996 Wiley-Liss, Inc.