IgM类抗HCV独特型抗体（抗HCV—Ab2）的ELISA法检测及意义

用抗人μ链单克隆抗体建立的ＥＬＩＳＡ法检测１５２例肝病患者ＩｇＭ类抗ＨＣＶ独特型抗体，甲肝阳性率０％（０／１５），乙肝８．５％（４／４７），丙肝３２．１％（２６／８１），丙肝合并乙肝者２２．２％（２／９）。丙肝组内频率为急肝０％（０／１７），慢性迁延性肝炎３８．１％（８／２１），慢性活动性肝炎４３．８％（１４／３２），肝炎肝硬化３６．４％（４／１１）。本法重复性好，批内变异系数４．６％，批间变异系数８．５％。用纯化人抗ＨＣＶ抗体进行阻断，阻断率＞８５％，丙肝组与乙肝组有显著性差异（Ｐ＜０．０１），与各种自身免疫性疾病无交叉，有较好的特异性和精密度，且与血清中ＩｇＭ抗ＨＣＶ关系密切，提示丙型肝炎ＩｇＭ类抗ＨＣＶ－Ａｂ２具有模拟抗原的作用，是慢性丙型肝炎的重要检测指标并与丙肝慢性化有关。     

石家庄地区新生儿尿巨细胞病毒感染的临床分析

采用聚合的酶链反应（ＰＣＲ）检测１０５例出生后３日内新生儿尿中巨细胞病毒,同时应用酶联免疫吸附法（ＥＬＩＳＡ）检测脐血特异性ＩｇＧ和ＩｇＭ抗体。结果,新生儿尿液ＨＣＭＶ－ＤＮＡ阳性者３３例,阳性率３１．４％,脐血ＩｇＧ抗体阳性率１００％,ＩｇＭ挤体阳性率１．９％。结果表明ＰＣＲ技术检测新生儿尿液ＨＣＭＶ－ＤＮＡ有利于ＨＣＭＶ先天性感染的早期诊断,早预防、早阻断。有利于更好地实行计划生育,有利于优生     

巨细胞病毒宫内感染的诊断及对胎儿的影响

采用地高辛探针斑点杂交技术和聚合酶链反应（ＰＣＲ）技术，对５１例人巨细胞病毒（ＨＣＭＶ）血清学抗体ＩｇＭ阳性孕妇的羊水、脐血和产后两周内的新生儿尿液，进行ＨＣＭＶＤＮＡ检测。同时与酶联免疫吸附法（ＥＬＩＳＡ）检测脐血、羊水中ＨＣＭＶ－ＩｇＭ的结果相比较。结果，地高辛探针斑点杂交检测ＨＣＭＶ－ＤＮＡ的阳性率分别为：羊水２１．５７％，脐血３３．３３％，新生儿尿液２７．４５％。ＰＣＲ检测ＨＣＭＶ－ＤＮＡ的阳性率分别为：羊水２９．４１％，脐血４３．１４％，新生儿尿液３Ｓ，２９％。羊水ＨＣＭＶ－ＩｇＭ阳性率为１１．７６％，脐血ＨＣＭＶ－ＩｇＭ阳性率为２３．５３％。脐血ＨＣＭＶ－ＤＮＡ阳性的新生儿平均出生体重明显低于脐血ＨＣＭＶ－ＤＮＡ阴性的新生儿，而脐血ＨＤＷ－ＤＮＡ阳性的新生儿肝功能异常、血小板减少以及低Ａｐｇａｒ评分的发生率明显高于脐血ＨＣＭＶ－ＤＮＡ阴性的新生儿。结果表明：采用ＥＲ技术检测羊水、脐血或产后两周内新生儿尿液ＨＣＭＶＤＮＡ有助于ＨＣＭＶ官内感染的早期诊断，便于临床早期干预，ＨＣＭＶ官内感染严重影响胎儿的生长发育，可造成胎儿肝功能异常、血小板减少和新生儿窒息的发生。     

丙型慢性肝病患者配偶中的HCV感染

丙型慢性肝病患者配偶中的ＨＣＶ感染［英］／ＡｋａｈａｎｅＹ…／／ＡｎｎＩｎｔｅｒｎＭｅｄ．－１９９４，１．２０（９）．－７４８～７５２为确定长期配偶ＨＣＶ感染的危险是否增加，作者对１５４例ＨＣＶ相关慢性肝病患者的配偶中的ＨＣＶ相关抗体和ＨＣＶＲＮＡ作...     

用聚合酶链反应和放射免疫法检测肾移植术后人巨细胞病毒感染

将５０例肾移植术后发热病人的尿标本接种细胞，用聚合酶链反应（ＰＣＲ）技术检测第１代细胞培养上清中的人巨细胞病毒（ＨＣＭＶ）ＤＮＡ。结果１９例为ＨＣＭＶＤＮＡ阳性，阳性率为３８．００％，用固相放射免疫法（ＲＩＡ）检测ＨＣＭＶ抗原，阳性的有１５例，阳性率为２７．７７％。将５４倒尿标本接种人胎肺二倍体单层细胞，分离到６株ＨＣＭＶ。用ＥＬＩＳＡ检测６５例病人单份血清的ＨＣＭＶ－ＩｇＭ抗体，阳性的有１７例，检测４５例病人双份血清ＩｇＧ抗体，１１例的第２份血清的ＨＣＭＶ－ＩｇＧ抗体滴度比第１份有４倍以上升高。上述结果表明，ＰＣＲ检测结果与ＲＩＡ的检测结果基本符合；病人血清抗体的阳性率与ＰＣＲ检测的阳性率基本符合。     

巨细胞病毒感染与可溶性白细胞介素2受体的关系

应用酶联免疫吸附试验（ＥＬＩＳＡ）对１０４例育龄妇女的血清进行了巨细胞病毒（ＨＣＭＶ）ＩｇＧ、ＩｇＭ抗体的检测，同时用ＥＬＩＳＡ双抗体夹心法测定了不同感染状态下血清中可溶性白细胞介素２受体（ｓＩＬ－２Ｒ）的水平，并将ｓＩＬ－２Ｒ水平与未感染ＨＣＭＶ的正常育龄妇女进行了比较。结果，育龄妇女中抗－ＨＣＭＶＩｇＧ的阳性率为８９．４％，ＩｇＭ的阳性率为９．６％，感染ＨＣＭＶ的妇女血清中ｓＩＬ－２Ｒ水平均大于未感染的对照组（１７８．１±５７．３Ｕ／ｍｌ），Ｐ＜０．０５，其中ＩｇＭ阳性者和ＩｇＭ、ＩｇＧ同时阳性者血清中ｓＩＬ－２Ｒ水平最高，分别为９１０±４６５．６Ｕ／ｍｌ和９０５±３４７．８Ｕ／ｍｌ，两者间的差异无显著性意义（Ｐ＞０．０５），但均大于仅抗－ＨＣＭＶＩｇＧ阳性者（４４６．８±１５８．９Ｕ／ｍｌ），Ｐ均＜０．０５。表明，ＨＣＭＶ感染可致ｓＩＬ－２Ｒ水平升高，并且活动性感染者上升明显。提示：ｓＩＬ－２Ｒ可能参与了ＨＣＭＶ的免疫致病机制。     

肿瘤患者巨细胞病毒感染及与输血,临床关系的研究

应用间接酶联免疫吸附试验检测了７０例肿瘤患者血清人巨细胞病毒特异性抗体，ＨＣＭＶ－ＩｇＭ和ＩｇＡ抗体的阳性率分别为６１．４２％和１４．２８％，明显高于对照组人群，恶性肿瘤组活动性ＨＣＭＶ感染率与良性肿瘤组相比有明显性差异。     

丙型肝炎病毒感染者中抗病毒IgM检测的意义

建立了抗ＨＣＶＩｇＭ间接ＥＬＩＳＡ方法，并用之检测ＨＣＶ不同感染人群。抗ＨＣＶＩｇＭ在不同感染人群中的检出率变化很大。急性输血后丙型肝炎患者检出率可高达９３．８％，而正常献血员中可低至０．６８％。比较抗ＨＣＶＩｇＭ和抗ＬｇＧ阳性中ＨＣＶＲＮＡ的检测结果发现，抗ＩｇＭ阳性者中，不同ＨＣＶ感染人群的ＨＣＶＲＮＡ检出率很高（９３．０％～１００％）；而ＩｇＧ阳性者中ＨＣＶＲＮＡ的检出率变化很大（３７．５％～９３．８％）。在４３例血液透析者中抗ＩｇＭ与ＨＣＶＲＮＡ检测的一致性为８３．７％；抗ＩｇＭ与抗ＩｇＧ检测的一致性为８８．４％，结果提示：（１）抗ＨＣＶＩｇＭ与ＨＣＶ活跃复制有关，（２）抗ＨＣＶＩｇＭ与抗ＨＣＶＩｇＧ检出不完全一致。因此，临床检测抗ＨＣＶＩｇＭ有其特殊意义。     

造血系统恶性肿瘤患者巨细胞病毒感染状态

造血系统恶性肿瘤患者巨细胞病毒感染状态作者采用核酸杂交方法对造血系统恶性肿瘤患者及正常人人类巨细胞病毒（ＨＣＭＶ）感染率进行了调查，其感染率分别为２８．６％和９．８％，二者差异显著。用间接ＥＬＩＳＡ方法测定血清中的ＨＣＭＶＩｇＭ抗体，发现患者组ＨＣＭ...     

成都地区孕妇和新生TORCH感染的检测与分析

采用ＥＬＩＳＡ检测孕妇血清及新生儿脐血ＴＯＲＣＨ特异性ＩｇＧ，ＩｇＭ抗体，同时采用ＰＣＲ检测ＣＭＶ－ＩｇＭ阳性孕妇的羊水及产后乳汁ＣＭＶ－ＤＮＡ，以了解孕妇及新生儿ＴＯＲＣＨ感染情况。结果显示ＴＯ，ＲＶ，ＣＭＶ，ＨＳＶ－１和ＨＳＶ－２检出率分别为４１．４８％，８９．３６％，９５．７４％，３９．３６％和２２．３４％，其活动性感染分别为４．２５％，３．１９％，２４．４６％，１１．７０％和５．３１％，新     

Antibody reactivity to human and vervet cytomegalovirus early antigen(s) in sera from patients with active cytomegalovirus infections and from asymptomatic donors.

Human fibroblasts were infected with vervet cytomegalovirus (VCMV) and cultured in medium containing 50 micrograms of cytosine arabinoside per ml for 72 h. Early antigens (EAg) were detected in the nuclei of infected cells by an indirect fluorescent antibody test with human sera having antibody to EAg of human cytomegalovirus (HCMV). Sera from patients with serological and/or virological evidence of active HCMV infection and from asymptomatic blood donors were examined for antibody activity with the HCMV and VCMV EAg's. The HCMV and VCMV EAg's were comparable in detecting levels of antibody activity and fluctuations in antibody titer of paired sera from patients. A total of 81% of patient sera reactive with HCMV EAg were also reactive with VCMV EAg. In contrast, only 14% of the asymptomatic donor sera reactive with HCMV EAg were also reactive with VCMV EAg. The VCMV EAg, therefore, appeared to differentiate latent from active infections in humans more effectively than did HCMV EAg.     

Evaluation of a microtiter plate fluorescent-antibody assay for rapid detection of human cytomegalovirus infection.

The use of monoclonal antibody (MAb) p63-27, which is reactive with the major immediate-early human cytomegalovirus (HCMV) protein pp72, was explored for the rapid diagnosis of HCMV viruria. The rapid assay detected all but 1 of 19 specimens identified by standard virus isolation methods from 1,676 newborn urine specimens, achieving a sensitivity of 94.5% and a specificity of 100%. The monoclonal antibody recognized 260 randomly obtained clinical isolates of HCMV, indicating the suitability of this reagent for use in screening assays. The sensitivity of the microtiter plate method declined rapidly for specimen from older infants and children with congenital CMV infection and virus-infected children attending a day-care center and was judged to be unacceptable for screening populations in this age group.     

Cytomegalovirus DNA detection in sera from patients with active cytomegalovirus infections.

Using a specific and sensitive polymerase chain reaction method, we detected reliably the presence of human cytomegalovirus (HCMV) DNA directly in serum samples collected at an early stage of HCMV infection, even before immunoglobulin M (IgM) antibodies were measurable. HCMV DNA was detected in serum from all patients with active HCMV infection; in 91% of these patients, HCMV DNA was found in the acute-phase serum. In 13 of 44 patients, HCMV DNA was found in serum before HCMV-specific IgM. For four kidney transplant recipients, the occurrence of HCMV DNA in serum, virus isolation from urine and leukocytes, and HCMV IgG and IgM serology were determined. We found a correlation between HCMV DNA in serum and positive virus isolation from leukocytes. In three of five congenitally infected infants, HCMV DNA and HCMV IgM were detected in the same sample. Two other infants were HCMV DNA positive, although no HCMV IgM antibodies were measurable. HCMV was found in urine from these infants either by virus isolation or with the polymerase chain reaction. Serum from one of the 22 healthy HCMV-seropositive blood donors was HCMV polymerase chain reaction positive.     

Diagnostic value of HCMV pp65 antigen detection by FCA for symptomatic and asymptomatic infection: compared to quantification of HCMV DNA and detection of IgM antibody in infants

Human cytomegalovirus (HCMV) can cause symptomatic or asymptomatic infection in infants. One hundred and twenty-six infants were assessed clinically for disease in infantile period. Eighty of them were classified as symptomatic infection on the basis of physical, instrumental, and laboratory findings, 5 were demonstrated by following up to have later developed HCMV disease, and the other 41 infants were classified as asymptomatic infection. HCMV DNA was positive in all urine samples of the symptomatic infants detected by quantitative polymerase chain reaction. HCMV-IgM antibody detected by chemiluminescent immunoassay (CLIA) was positive in 62 of the 85 symptomatic infants, but was negative in all of the samples of asymptomatic infants. HCMV pp65 antigen detected by flow cytometry assay (FCA) was positive in 77 of the 85 symptomatic infants and in none of the asymptomatic infants. The coincidence to symptom of HCMV pp65 antigen detection was higher than those of HCMV DNA and HCMV-IgM antibody detection. The sensitivity, specificity, positive prognostic value and the negative prognostic value of HCMV pp65 antigen detection for diagnosis of HCMV infection was 90.6, 100, 100 and 83.7%, respectively. We concluded that detection of pp65 antigen by FCA is more sensitive for diagnosis of HCMV infection than detection of HCMV-IgM antibody and is better than HCMV DNA quantification for distinguishing the symptomatic and asymptomatic HCMV infection in infants.     

尿毒症患者巨细胞病毒感染及与输血关系的研究

应用间接酶联免疫吸附试验法检测５２例尿毒症患者血清中巨细胞病毒特异性ＩｇＭ，ＩｇＡ抗体。结果表明特异性ＩｇＭ，ＩｇＡ抗体阳性率分别为６９．６％和１３．８％，明显高出对照组人群，统计学处理Ｐ〈０．０１，差异非常显著。     

Detection of human Betaherpesvirinae in saliva and urine from immunocompromised and immunocompetent subjects.

Human cytomegalovirus (HCMV) is a well-known opportunistic agent that reactivates in human immunodeficiency virus (HIV)-seropositive subjects. Human herpesvirus 6 (HHV-6) and HHV-7 were discovered recently and, like HCMV, belong to the Betaherpesvirinae subfamily. We looked for the presence of HCMV, HHV-6, and HHV-7 by PCR with saliva and urine samples from 125 HIV-seropositive patients at different stages of HIV infection and with saliva and urine samples from 29 HIV-seronegative subjects. All three viruses were frequently detected in the saliva (overall rates of detection, 61, 43, and 63% for HCMV, HHV-6, and HHV-7, respectively) with no correlation with the stage of immune deficiency. In contrast, HCMV was detected in urine much more frequently than the two other herpesviruses (overall rates of detection, 37, 2, and 6.5% for HCMV, HHV-6, and HHV-7, respectively) and was associated with immune deficiency. This suggests that these three genetically related viruses differ from each other with regard to replication in the urinary tract.     

Human cytomegalovirus (HCMV)-specific immunoglobulin E as a serologic marker for HCMV infection in immunocompromised patients

Summary An antibody capture assay using an enzyme-linked human cytomegalovirus (HCMV) antigen for the detection of specific immunoglobulin E (IgE) was established. IgG, M, and E responses to HCMV were studied in 497 sera obtained from 44 renal transplant recipients and 51 acquired immunodeficiency syndrome (AIDS) patients. The results were compared with those obtained from 58 HCMV-seropositive healthy individuals. HCMV-specific IgE was detected in 11 (91.7%) renal transplant recipients with primary HCMV infection. In contrast, antibodies of the IgG and IgM classes were detected in only 6 (50.0%) of these patients. Specific IgE was detected in 10 (90.9%) out of 11 renal allograft recipients suffering from secondary HCMV infection. Significant IgG titer rises and IgM were detected in 2 (18.2%) and 6 (54.6%) of these patients, respectively. IgG titer rises and IgM and IgE antibodies were seen in 5 (12.2%), 1 (2.4%) and 18 (43.9%) AIDS patients respectively. All healthy immunocompetent HCMV-seropositive individuals were tested IgE negative. The results obtained in our study indicate that IgE against HCMV is a more reliable serologic marker for primary and secondary HCMV infection than IgM in immunocompromised individuals, especially in organ transplant recipients, since it is not affected by the prophylactic application of HCMV hyperimmune globulin preparations.Abbreviations AIDS acquired immunodeficiency syndrome - BAL bronchoalveolar lavage - CDC Centers for Disease Control, Atlanta, USA - ELISA enzyme-linked immunosorbent assay - HCMV human cytomegalovirus - HIV human immunodeficiency virus - Ig immunoglobulin - PBS phosphate buffered saline - RTR renal transplant recipients     

Direct detection of human cytomegalovirus in urine specimens from renal transplant patients following polymerase chain reaction amplification

A polymerase chain reaction (PCR) assay was used to amplify human cytomegalovirus (HCMV) directly from urine specimens taken from renal transplant patients. In serial urine samples from patients who had at least one specimen positive for HCMV; the PCR assay consistently detected the presence of HCMV DNA sequences, whereas virus detection by other tests such as enzyme-linked immunosorbent assay (ELISA), nonradioactive DNA hybridization assay, and virus isolation were variable. Of 37 specimens positive by PCR, 36 were positive by either ELISA, hybridization assay, or virus isolation. Infectious virus was detected in 13 of the 37 PCR-positive urines. HCMV DNA was detected by PCR in all samples that were positive for HCMV by either hybridization assay or virus isolation. The viral genome copy number was determined by PCR assay for several urine samples that were positive by virus isolation but negative for HCMV by ELISA or hybridization assay. Viral genome copy number estimates indicated the presence of HCMV at very low levels in these urines verifying the fidelity of the virus isolation procedures. The consistency of the PCR assay makes it an ideal method for detection of infection and monitoring antiviral drug therapy in patients infected with HCMV.     

Evaluation of antigen and antibody detection in urine specimens from children with congenital human cytomegalovirus infection

Fetal infection with human cytomegalovirus (HCMV) is the leading viral cause of brain dam age among newborns at birth or later in life. Efforts to screen newborns routinely for shedding of the virus by immunoassay have been ham pered by inhibitors in urine, reportedly the host protein beta2-microglobulin (β2m). An enzyme-linked immunosorbent assay (ELISA) was devel oped for the detection of HCMV antigen in which the reactivity was not affected by the presence of β2m, but nevertheless inhibition was observed when urine samples with high levels of virus were tested. The presence of antibodies to HCMV was demonstrated in these urine samples by antibody ELISA and immunoblot using the major antigenic protein of HCMV (pp150) expressed in Escherichia coli; this offers an alterna tive explanation for the inhibition in ELISA. The presence of HCMV antibodies correlated significantly with congenital HCMV infection (as detected by tissue culture isolation of virus from urine samples of newborns), especially with as ymptomatic cases (sensitivity 70%; specificity 94%). The data indicate a local (renal) immune response to HCMV in congenitally infected children, which may have future diagnostic applica tions. © 1995 Wiley-Liss, Inc.

1 This article is a US Government work and, as such, is in the public domain in the United States of America.

     

The use of ELISA and nonradioactive DNA hybridization assays for the detection of human cytomegalovirus

A hybridization assay using a biotinylated DNA probe was compared to both ELISA and direct isolation methods for detecting human cytomegalovirus (HCMV). The biotin labeled HCMV AD 169 HindIII-O-DNA fragment was used in a dot-blot assay to screen for the presence of HCMV in 186 urine specimens obtained from kidney transplant patients. The biotinylated HCMV HindIII-O probe could detect 3 log10 TCID50 units of HCMV. Urine specimens were also examined for the presence of HCMV by either ELISA or direct isolation of virus in tissue culture. The HindIII-O fragment detected 12 of 20 culture positive samples (sensitivity, 60%). There were 5 samples which were probe positive and cell culture negative (specificity, 97%). The ELISA assay also detected 12 of 20 culture positive samples (sensitivity, 60%). Eight samples were ELISA positive, cell culture negative (specificity, 95%). Seven specimens were positive by all three criteria. Five specimens which were both ELISA positive and probe positive were cell culture negative. The ELISA positive, probe positive, culture negative specimens originated from patients who gave a culture positive specimen within 10 days of the original sample. The combination of probe and ELISA assays detected 16 of the 20 culture positive specimens (sensitivity, 80%). The combined use of biotinylated DNA probes and ELISA allows the detection of HCMV in urine specimens with good sensitivity and specificity.