狂犬病毒新型快速荧光斑点抑制实验方法的建立

目的建立一种新型的快速荧光斑点抑制实验用于狂犬病毒抗体的检测。方法本研究以携带绿色荧光蛋白基因的嵌合狂犬病毒HEP-GFP株为基础毒株,建立了一种新型快速荧光斑点抑制实验(RFFIT-GFP);并对多份人、犬、猫的血清进行了检测,还将该检测结果与传统的RFFIT,ELISA相比较。结果与结论RFFIT-GFP和RFFIT,ELISA测定的结果基本相一致,特异性都比较高,而且RFFIT-GFP是一种比它们更为简便、经济的检测方法。     

不同HCV抗体检测试剂效果的比较

目的对不同种类的实验室HCV抗体检测试剂进行比较。方法选取天门市第一人民医院采用国内实验室常用的HCV抗体筛查试剂(科华﹑新创﹑万泰和雅培)检测呈阳性的205份样本作为研究对象,进行HCV RNA核酸扩增检测(NAT),若NAT检测为阴性,再进行重组免疫印迹实验(RIBA)进一步确认。对4种筛查试剂的检测结果进行比较,并对其S/CO值和真阳性率进行分析。结果任一筛查试剂检测结果呈阳性反应的205份样本中,4种筛查试剂均为阳性的有191份(93.2%),4种筛查试剂结果不一致的有14份(6.8%)。新创、科华、万泰及雅培试剂的阳性预测值分别为88.2%(180/204)、93.8%(180/192)、91.4%(180/197)及90.0%(180/200)。筛查试剂阳性预测值≥95%的S/CO阈值分别为:新创9.0、科华4.0、万泰5.0及雅培7.0。结论科华、新创、万泰和雅培4种筛查试剂阳性预测值≥95%的S/CO阈值之间相差较大,与国内其他实验室比较相差也较大。当前各实验室应建立适用于自身的阳性预测值≥95%的S/CO阈值,以减少确证试验样本数量。     

Neutralizing antibody response to hepatitis C virus

A critical first step in a "rational vaccine design" approach for hepatitis C virus (HCV) is to identify the most relevant mechanisms of immune protection. Emerging evidence provides support for a protective role of virus neutralizing antibodies, and the ability of the B cell response to modify the course of acute HCV infection. This has been made possible by the development of in vitro cell culture models, based on HCV retroviral pseudotype particles expressing E1E2 and infectious cell culture-derived HCV virions, and small animal models that are robust tools in studies of antibody-mediated virus neutralization. This review is focused on the immunogenic determinants on the E2 glycoprotein mediating virus neutralization and the pathways in which the virus is able to escape from immune containment. Encouraging findings from recent studies provide support for the existence of broadly neutralization antibodies that are not associated with virus escape. The identification of conserved epitopes mediating virus neutralization that are not associated with virus escape will facilitate the design of a vaccine immunogen capable of eliciting broadly neutralizing antibodies against this highly diverse virus.     

Immunoglobulin M antibody to hepatitis C virus during interferon therapy for chronic hepatitis C.

Testing for immunoglobulin (Ig) M antibody to hepatitis C virus (anti-HCV) as a predictive factor of therapeutic response to recombinant interferon alfa (rIFN-alpha) in chronic hepatitis C was evaluated in 122 patients with IgG anti-HCV. IgM anti-HCV was present in the pretreatment sample of 88% of patients who responded to treatment, including 20 of 21 (95%) long-term responders and 24 of 29 (83%) responders who had relapses after cessation of therapy. In contrast, IgM anti-HCV was present in only 23 of 39 (59%) nonresponders and 22 of 33 (66%) untreated controls (P less than 0.05). The number of cases with detectable IgM anti-HCV tended to decrease in responder patients, which was more evident for complete responders (42%) than for responders who relapsed (72%). During follow-up, the antibody became undetectable in the majority of long-term responders (28% were still IgM anti-HCV positive) but remained detectable in 69% of responders who relapsed (P less than 0.05). No special changes were noted in nonresponder or control patients. Thus, testing for IgM anti-HCV may help to identify a subset of patients who will benefit from rIFN-alpha therapy in chronic hepatitis C.     

Peroxidase-labeled antibody technique for rapid detection of mouse hepatitis virus in cases of natural outbreaks.

A recent outbreak of hepatitis in mice at the Laboratory Animals Centre (Surrey, England) is described. The only strains of mice that showed clinical symptoms and typical liver lesions were the nude (nu/nu) and obese mice. Although other inbred strains showed no clinical symptoms, all strains seroconverted. Neutralization typing of the mouse hepatitis virus showed it to be type 1, a hepatotropic strain of low virulence, a finding that was consistent with the pathology of the disease. Because of recurring outbreaks of disease caused by this strain and other strains of MHV in the United Kingdom, a rapid method for diagnosis in tissue culture was devised, employing the peroxidase enzyme-labeled antibody technique for detection of mouse hepatitis virus in L-929 cells.     

IgM antibody to hepatitis C virus in acute and chronic hepatitis C.

To assess possible role of testing for IgM-specific antibody in the diagnosis and monitoring of patients with hepatitis C, we tested sera from 14 patients with acute and 97 patients with chronic non-A, non-B hepatitis for IgG and IgM antibody to hepatitis C virus. IgG antibody to hepatitis C virus was detected in 93% of acute cases and 91% of chronic cases. Of the 101 patients with IgG antibody to hepatitis C virus, 57% had IgM antibody to hepatitis C virus. None of the 20 healthy subjects or 40 patients with acute or chronic hepatitis A or hepatitis B had IgM antibody to hepatitis C virus. At the onset of clinical symptoms in acute hepatitis C, IgG antibody to hepatitis C virus was detected in 8 (57%) and IgM antibody to hepatitis C virus in 9 of 14 patients (64%). Eventually, both IgG and IgM antibody to hepatitis C virus became detectable in 13 of 14 patients with acute hepatitis C. Seven patients with antibody to hepatitis C virus resolved the acute infection within 6 mo and all seven cleared IgM antibody to hepatitis C virus, whereas two cleared IgG antibody to hepatitis C virus. Six patients had a chronic outcome of the acute infection and IgM antibody to hepatitis C virus persisted in detectable amounts for more than 6 mo in all (mean = 15.5 mo). Among 88 patients with chronic non-A, non-B hepatitis with IgG antibody to hepatitis C virus, IgM antibody to hepatitis C virus was detected in 45 (51%).(ABSTRACT TRUNCATED AT 250 WORDS)     

A new enzyme immunoassay for the detection of antibody to hepatitis E virus

Background and Aim : The purpose of the present study was to develop enzyme immunoassay (EIA) for the detection of IgG anti‐hepatitis E virus (HEV) activity using two new recombinant proteins as antigenic targets, and to evaluate these EIA with the aid of statistical methods. Methods : Two proteins, a mosaic protein and pB166 containing region 452–617 aa of the ORF2 of the HEV Burma strain, were used to develop the new HEV EIA. This EIA was evaluated using several panels of serum specimens obtained from: (i) acutely HEV‐infected patients; (ii) patients with non‐A, non‐C hepatitis; (iii) normal blood donors (NBD) from non‐endemic countries; and (iv) experimentally infected chimpanzees. Results : A new HEV EIA was developed using two new recombinant proteins. This assay was able to detect anti‐HEV activity in all specimens from acutely HEV‐infected patients. When NBD were tested, more than 15% of specimens were found to be IgG anti‐HEV positive. All NBD anti‐HEV‐positive specimens were tested with overlapping synthetic peptides spanning the entire HEV ORF2‐encoded protein. More than 90% of the anti‐HEV‐positive NBD specimens immunoreacted with an average of 15 synthetic peptides derived from different regions of the HEV ORF2 protein. These data suggest that the HEV EIA is at least 90% specific in detecting remote HEV infections. Conclusion : The new HEV EIA developed in the present study is a highly specific diagnostic assay for the detection of anti‐HEV activity in serum specimens obtained from different epidemiologic settings. © 2002 Blackwell Publishing Asia Pty Ltd     

The significance of antibody to hepatitis C virus in patients with chronic hepatitis B.

We assessed the prevalence and clinical significance of antibodies to hepatitis C virus among a cohort of 148 patients with chronic hepatitis B virus infection. Sixteen patients (11%) had anti-hepatitis C virus detectable by enzyme-linked immunoassay. The results from eight of these patients were positive by recombinant immunoblot assay. The results of recombinant immunoblot assay testing were not consistent; therefore the analysis of the patients' data was based on anti-hepatitis C virus enzyme-linked immunoassay results. Patients with chronic hepatitis B with anti-hepatitis C virus were more likely to be cirrhotic (44% vs. 21%) and to have decompensated liver disease (24% vs. 6%). Hepatitis B virus replication appeared to be suppressed in patients with both infections as measured by hepatitis B virus-associated DNA polymerase activity (mean = 2,055 vs. 2,555 cpm). Human immunodeficiency virus infection was more common in the anti-hepatitis C virus positive group (36% vs. 11%). Thus hepatitis C virus appears to suppress hepatitis B virus replication and to cause more severe liver disease in patients with chronic hepatitis B infection.     

Significance of IgM antibody to hepatitis C virus in patients with chronic hepatitis C.

We assessed the correlation between the positivity for serum IgM antibody to hepatitis C virus and the activity of liver disease in patients with chronic hepatitis C virus infection. Serum samples were taken from 10 antibody to hepatitis C virus-positive asymptomatic patients with normal serum ALT levels, from 14 untreated patients with clinically and histologically proven chronic hepatitis C and from 26 patients with clinically and histologically proven chronic hepatitis C assigned to receive recombinant interferon alpha-2a (6 million IU three times a week for 6 mo). Each serum specimen was tested for IgM antibody to hepatitis C virus-associated C100-3 antigen by enzyme-linked immunosorbent assay. Patients were observed for at least 12 mo. All 10 patients with normal ALT values tested negative for IgM antibody to hepatitis C virus. In contrast, 33 of 40 (82%) patients with chronic hepatitis C had IgM antibody to hepatitis C virus, and a positive correlation was seen between the ALT level and the level of IgM antibody to hepatitis C virus (r = 0.803, p less than 0.001). During interferon treatment, ALT levels declined into the normal range in 18 of 26 treated patients (69%) and remained normal after stopping treatment in 8 patients (31%). In untreated patients, in treated patients who did not respond to interferon treatment and in responder patients who relapsed, no significant changes in IgM antibody to hepatitis C virus levels were seen during the study period. In contrast, IgM antibody to hepatitis C virus became undetectable by the end of interferon treatment in seven of eight patients with a sustained response.(ABSTRACT TRUNCATED AT 250 WORDS)     

The epidemiology of hepatitis C virus antibody in Yemen.

A cross-sectional survey of 348 subjects without evidence of liver disease was conducted to investigate the prevalence and risk factors for hepatitis C virus antibody (anti-HCV) seropositivity in the Yemen Arab Republic. The mean age of study subjects was 28.7 years (range 3-80), and 61% were males. Using commercial enzyme-linked immunosorbent assays (ELISA), 6.0% (95% confidence interval [CI] 3.8-9.1) of subjects were anti-HCV-positive, 13.5% were hepatitis B surface antigen-positive (HBsAg-positive), and 51.4% were positive for at least one serologic marker of prior hepatitis B infection. Nine (2.6%; 95% CI 1.2-4.9) of the 21 ELISA-positive sera were confirmed to be anti-HCV positive by a recombinant immunoblot assay. Anti-HCV seropositivity was significantly associated with age (odds ratio [OR] 2.0 for each 10-year increase in age) and prior surgery (OR 10.1), but was not associated with a history of prior blood transfusion or markers of hepatitis B infection. These preliminary data suggest that hepatitis C may pose a substantial health threat in Yemen.     

Maternal neutralizing antibody and transmission of hepatitis C virus to infants

To determine whether lower levels of hepatitis C virus (HCV)-specific neutralizing antibodies (nAb) are associated with an increased risk of mother-to-child transmission (MTCT) of HCV, HCV nAb titers were assessed in 63 mothers coinfected with HCV and human immunodeficiency virus (HIV) type 1. Of the mothers, 16 transmitted HCV to their infant, but no difference was detected between the ability of maternal plasma from transmitters and nontransmitters to neutralize heterologous HCV pseudoparticles (median nAb titer, 1:125 vs. 1:100; P = .23). In the setting of HIV/HCV coinfection, we found no evidence that HCV nAbs are associated with the prevention of MTCT of HCV.     

Performance characteristics of a combined hepatitis C virus core antigen and anti-hepatitis C virus antibody test in different patient groups

We evaluated the performance of a hepatitis C virus (HCV) antigen/antibody combination test [Murex HCV Antigen/Antibody Combination Test (Murex Ag/Ab test)] by comparing it with the current third-generation HCV antibody enzyme immunoassay (anti-HCV). A total of 403 serum samples were consecutively collected from four patient groups: healthy controls (n=100); HCV-infected patients (HCV?group, n=102); Human immunodeficiency virus (HIV)/HCV-infected patients (HIV/HCV group, n=100); and patients with uremia (uremia group, n=101). Performances were evaluated for the Murex Ag/Ab, anti-HCV, and HCV RNA in the HIV/HCV and uremia patient groups. In the HCV group, all 102 samples showed concordant positive and negative results for anti-HCV, Murex Ag/Ab, and HCV RNA tests. In the HIV/HCV group, all 100 samples were positive for both anti-HCV and Murex Ag/Ab tests, whereas 88 patients (88%) were HCV RNA positive. In the uremia group, 14 (69.0%) of the 23 anti-HCV-positive patients were HCV RNA positive, whereas 14 (77.8%) of the 18 Murex Ag/Ab-positive patients were HCV RNA positive. None of anti-HCV-negative or Murex Ag/Ab-negative patients were HCV RNA positive. Based on the HCV RNA assay, the sensitivities for both anti-HCV and Murex Ag/Ab assays were 100%, whereas the specificities of these two assays were 89.7% and 95.4%, respectively. With good sensitivity and specificity, the Murex Ag/Ab assay could be a useful alternative diagnostic tool, especially in immunocompromised populations, such as patients with uremia or those infected with HIV.     

Frequency of antibody to hepatitis C virus in asymptomatic HBsAg-negative chronic active hepatitis.

To determine the frequency of antibodies to hepatitis C virus in asymptomatic patients with HBsAg-negative chronic active hepatitis, sera from 30 consecutive patients with few or no symptoms of liver disease were tested by an enzyme immunoassay. The reactivity of antibodies detected by enzyme immunoassay against hepatitis C virus encoded antigens was determined by recombinant immunoblot assay. Antibodies were detected in 11 of the 30 patients (37%) and eight of the seropositive sera (73%) were reactive by recombinant immunoblot assay. Nonreactive patients were weakly positive by enzyme immunoassay (sample/cutoff ratio, less than or equal to 1.9) in contrast to reactive patients (sample/cutoff ratio, greater than or equal to 6.3). The prevalence of immunoserologic markers was similar in patients with and without antibodies (78 vs. 87%) but high titers (greater than or equal to 1:160) were more common in seronegative patients (53 vs. 11%). Additionally, seronegative patients had smooth muscle antibodies (83 vs. 25%, p less than 0.05) and concurrent extrahepatic immunologic diseases (37 vs. 9%) more commonly than seropositive counterparts. We conclude that asymptomatic patients with HBsAg-negative chronic active hepatitis frequently have antibodies to hepatitis C virus. These antibodies commonly react to specific viral antigens, especially if the enzyme immunoassay is strongly positive. Seropositive patients infrequently have concurrent immunologic disorders or smooth muscle antibodies. Immunoserologic markers lack diagnostic specificity except in higher titer.