Quantitative assay for anti-HCV core antibody in healthy individuals found anti-HCV positive during a mass screening

We investigated the relationship between the viremia of hepatitis C virus (HCV) and anti-HCV-core antibody (anti-C11) titer in 20 individuals found to have second-generation HCV antibody during a mass screening for liver disease. Of those 20, anti-C11 titer was less than 10 units in six (30%), and higher than 10 units in 14 (70%). Serum HCV-RNA was not detected in the six individuals in the lower titer group, as compared with 13 (93%) of the 14 individuals in the higher titer group (P < 0.001). Similarly, abnormalities in the zinc sulfate turbidity test (ZTT) were significantly more frequent in the higher titer vs. the lower titer group (71.4% vs. 0%;P < 0.01). Of 248 patients with chronic liver disease, only five (2%) patients had anti-C11 titers below 10 units. Thus, the anti-HCV-core antibody titer was closely related to the status of HCV replication, and is useful for identifying individuals with or without active replication of HCV.     

An inter-laboratory study of anti-HCV antibody detection in saliva samples

AIM: The purpose of this study was to evaluate the efficacy of anti-hepatitis C virus (HCV) antibody detection in the saliva samples of 108 drug users in an inter-laboratory study. METHODS: Between January and June 2001, 108 subjects in Lille, Metz and Lens received a test to detect anti-HCV antibodies in their saliva. Two consecutive saliva samples were taken in each subject (Salivette system, Sarstedt). An HCV serology (Axsym HCV 3.0, Abbott) was also performed and serum HCV RNA detection by Amplicor HCV 2.0 (Roche) was performed when HCV serology was positive. Sixty three patients had a negative HCV serology, 45 had a positive HCV serology, and 31 of these had positive HCV RNA as well. Tests for the detection of the anti HCV antibody in saliva samples were performed as a blind study in both the Lille and the Thionville laboratories. RESULTS: The sensitivity of saliva anti-HCV antibody tests was respectively 71% (32/45) and 78% (35/45) in Lille and Thionville. In the event of positive HCV viremia, the sensitivity was respectively 90% (28/31) and 93% (29/31). The specificity was respectively 97% (61/63) and 98.5% (62/63). Results from the two laboratories agreed for 101 saliva tests while discrepancies were found in 7 (Kappa Concordance Coefficient: 0.85). CONCLUSIONS: This study confirms, in a large, unselected population sample, that anti-HCV antibody detection tests in saliva allow the detection of 90% of viremic HCV-antibody-positive patients with excellent specificity. The simplicity and reproductibility of this technique makes it a precious tool for epidemiological studies.     

Epidemiological, biological and histological characterization of patients with indeterminate third-generation recombinant immunoblot assay antibody results for hepatitis C virus

We studied the epidemiological, laboratory and histological characteristics of a group of patients with positive antibodies against hepatitis C virus (HCV) as determined by third-generation enzyme-linked immunosorbent assay (ELISA), and with indeterminate HCV antibody positivity as established by third-generation recombinant immunoblot assay (RIBA-3). The results obtained were compared with those recorded in a group of RIBA-3-positive patients. Both groups correspond to blood donors in whom the prevalence of hepatitis C is low. There were no statistically significant intergroup differences in mean age, or in the presence of infection risk factors. RNA positivity was much more frequent in the RIBA-positive group (71%vs 10%; P < 0.05), as was transaminase elevation during the 3 years of follow-up (54%vs 13%; P < 0.05). In 46% of the RIBA-indeterminate patients the liver biopsy proved normal, or only liver steatosis or minimal changes were detected, while 33% had persistent chronic hepatitis, and 21% showed active chronic hepatitis. A mean Knodell index score of 2.28 was recorded; 50% of the subjects showed no fibrosis, 46% grade 1 fibrosis (fibrous portal expansion), 4% grade 2 fibrosis (bridging fibrosis), and none grade 3 fibrosis (liver cirrhosis). In the RIBA-positive group, a greater percentage of patients had active chronic hepatitis, a greater Knodell index, and increased-grade fibrosis. It can be concluded that the RIBA-3-indeterminate group is epidemiologically similar to the RIBA-3-positive series, although with a lesser prevalence of laboratory test alterations, a lower viral replication index, and more likely to have benign disease - particularly in subjects without viral replication.     

Preliminary study of a dot immunogold filtration assay for rapid detection of anti-HCV IgG

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Low prevalence of anti-HCV antibody among Italian children

Summary The seroprevalence of anti-HCV antibody was studied among 2,749 children and teenagers (1,438 males and 1,311 females) living in Italy. Anti-HCV antibody testing was positive by both EIA and RIBA in ten (0.36%) subjects. The positivity rate increased with age, ranging from 0 among children less than 6 years of age to 0.8% among those aged 17–19 years x² linear regression=0.038). Anti-HCV prevalence ranged from 0.2% in northeastern regions and in Apulia to 0.6% in Sicily and Sardinia (p>0.005), and no difference was seen between males (0.35%, C.I. 95%: 0.04–0.66) and females (0.38%, C.I. 95%:0.04–0.66) (Fisher's exact test=0.565). From these data it appears that in Italy HCV infection is an uncommon event during childhood.

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Niedrige Prävalenz von Anti-HCV-Antikörpern bei italienischen Kindern

Zusammenfassung Bei 2.749 Kindern und Jugendlichen (1.438 Jungen und 1.311 Mädchen), die in Italien leben, wurde eine Studie zur Seroprävalenz der anti-HCV Antikörper durchgeführt. Bei zehn der Getesteten (0,36%) fand sich mit EIA und RIBA ein positiver Befund. Die Rate an positiven Fällen nahm mit dem Alter zu von 0 bei Kindern unter 6 Jahren auf 0,8% bei den 17–19jährigen (Chi² lineare Regression=0,038). In den nordöstlichen Regionen and Apulien lag die anti-HCV Seroprävalenz bei 0,2%, in Sizilien und Sardinien bei 0,6% (p>0,005). Zwischen Mädchen und Jungen fand sich kein Unterschied (0,35%, 95% CI: 0,04–0,66 bei Jungen und 0,38%, 95% CI: 0,04–0,66; Fisher's exakter Test 0,565). Aus diesen Daten läßt sich ableiten, daß die HCV-Infektion in der Kindheit in Italien ein seltenes Ereignis ist.

     

Serodiagnosis of neurocysticercosis in patients with epileptic seizure using ELISA and immunoblot assay

Sera from 88 patients from Santa Catarina and S?o Paulo states of Brazil, with epileptic seizures who underwent cerebral computed tomography (CT) were analyzed for the detection of antibodies to T. solium cysticercus by ELISA and Immunoblot (IB) with the following antigens: Taenia solium cysticercus total saline (Tso), Taenia crassiceps cysticercus vesicular fluid (Tcra-vf) and T. crassiceps cysticercus glycoproteins (Tcra-gp). ELISA carried out with Tso, Tcra-vf and Tcra-gp antigens showed 95%, 90% and 80% sensitivities, respectively, and 68%, 85% and 93% specificities, respectively. In the epileptic patients group, ELISA positivity was 30%, 51% and 35% with Tso, Tcra-vf and Tcra-gp antigens respectively. Considering the IB as the confirmatory test, the positivity was 16% (14/88) in the epileptic patients total group and 22% (12/54) in the epileptic patients with positive CT and signals of cysticercosis. We found a significant statistical correlation among ELISA or IB results and the phase of the disease when any antigens were used (p < 0.05). We emphasize the need to introduce in the laboratory routine the search for neurocysticercosis (NC) in patients presenting with epileptic seizures because of the high risk of acquiring NC in our region and its potential cause of epilepsy.     

Prevalence of antibody to Borrelia burgdorferi by indirect fluorescent antibody assay, ELISA, and western immunoblot in healthy adults in Wisconsin and Arizona.

The prevalence of antibody to Borrelia burgdorferi in healthy adults from Wisconsin and Arizona was determined by indirect fluorescent antibody assay (IFA), ELISA, and Western immunoblotting. A total of 301 sera from adult volunteer blood donors were collected from three areas of Wisconsin and compared with 49 consecutive anonymous adult volunteer donor sera from Tucson, Arizona, an area without reported Lyme borreliosis. Regional differences in seropositivity were found for Western immunoblotting (34[11%] of 301 from Wisconsin and none of 49 from Tucson; P less than .01) but not IFA or ELISA. No correlation was found among Western immunoblotting and IFA or ELISA results. For persons living in Madison or Milwaukee, Wisconsin (cities not endemic for Lyme borreliosis), 19 (86%) of 22 with a positive Western blot, but only 12 (48%) of 25 with a positive IFA or ELISA, had a significant exposure risk to B. burgdorferi-infected Ixodes dammini (odds ratio, 6.9; 95% confidence interval, 1.4-43.5). Western blot results were consistent with epidemiologic exposure to B. burgdorferi and implied frequent asymptomatic infection among healthy adults living in or visiting areas endemic for Lyme borreliosis.     

HIV-2 antibody testing of blood donors with doubtful immunoblot results for HIV-1

Summary Antibodies against human immunodeficiency virus type-1 (HIV-1) in samples from blood donors are commonly detected by various enzyme-linked immunosorbent assays (ELISA) and by confirmatory tests, e.g., Western blot or immunofluorescence tests. Immunoblot reactivity, which is directed only towards the HIV-1 core proteins p 18, p 24 and p 55, may represent false-positive reactions. Out of 125,000 blood donations, 140 were repeatably HIV-1 antibody reactive by ELISA; of these, 20 were doubtful positive sera with isolated p 18 and/or p24 bands in the HIV-1 confirmatory assay. Antibodies to HIV-2 are known to cross-react with these HIV-1 core proteins. We therefore assayed the 20 sera by immunofluorescence and immunoblotting for the presence of antibodies to HIV-2. None of these doubtful HIV-1 antibody positive blood donor sera was found to have antibodies to HIV-2.     

Hepatitis C antibody assay in a longitudinal study of haemophiliacs

Stored sera from 28 patients with inherited coagulation disorders who had developed non-A non-B hepatitis (NANBH) following a first exposure to clotting factor concentrates and 15 similar, but unmatched, patients who had received blood products but had normal transaminases on sequential testing were tested using the Ortho enzyme-linked immunosorbent assay (ELISA) anti-HCV assay. Twenty-seven of the 28 patients with NANBH were anti-HCV positive after exposure. In 10 of those in whom dates of first exposure and seroconversion were well-defined, the median time interval to NANBH was 4 weeks (range 1-7) and to anti-HCV seroconversion was 11 weeks (range 7.5-14.5). None of the 15 patients without NANBH developed anti-HCV. This first generation Ortho ELISA anti-HCV assay showed 96% sensitivity and 100% specificity and has potential use as an adjunct in the surveillance of new clotting factor products.     

Evaluation of a third generation anti-HCV assay in predicting viremia in patients with positive HCV antibodies

The most practical screening test for hepatitis C virus antibodies are second and third- generation enzyme immunoassays. We evaluated the usefulness of the third generation microparticle enzyme immunoassay (MEIA) in predicting HCV viraemia in anti-HCV positive patients. Serum samples from 106 patients with positive anti-HCV were obtained. To evaluate the diagnostic value of the MEIA test in predicting HCV viraemia, anti-HCV positive patients were categorized in two groups according to the presence or absence of serum HCV-RNA. Among the 106 patients, 26 had non detectable serum HCV-RNA and 80 had detectable HCV-RNA by PCR. The assay automatically calculates a result based on the ratio of sample rate to the cut-of rate for each sample and control (S/CO). When the means of S/CO values for patients with detectable and non detectable HCV-RNA were analyzed, a statistically significant difference was found, (79.3 SD 22.2 vs. 8.2 SD 6.4, respectively) (p 0.0001). We further analyzed the best cut-off value of the S/CO in differentiating viremic from non viremic patients. The S/CO value of 26 showed a sensitivity of 99% and a specificity off 96% in discriminating both categories of HCV infected patients. In conclusion, our data demonstrate that viremic HCV patients had higher S/CO values in the MEIA test in comparison with non viremic patients. Hence, this assay may be used to predict HCV viraemia in anti-HCV positive individuals.     

Indeterminate third-generation hepatitis C recombinant immunoblot assay and HCV RNA analysis: isolated reactivity against NS5 associated with HCV viraemia in clinical patients but not blood donors

It has previously been reported that singular reactivity against NS5 in the third generation hepatitis C virus (HCV) confirmatory recombinant immunoblot assay (RIBA-3) is not associated with detectable HCV RNA in sera. In order to investigate the significance of indeterminate HCV RIBA-3 results with particular regard to NS5, 165 sera with indeterminate RIBA-3 results were analysed for the presence of HCV RNA. Sera from blood donors constituted 58 of the 165 samples, whereas 11 were from immunocompromized patients. The remaining 96 sera were from non-immunosuppressed clinical patients. Of the 107 RIBA-3 indeterminate samples from clinical patients, 19 were HCV RNA positive (18%), with 3 of the 57 (5%) NS5 reactive samples having detectable HCV RNA. None of the 58 indeterminate samples obtained from blood donors had detectable HCV RNA. Thus, having an indeterminate RIBA-3, including singular reactivity against NS5, may be associated with the presence of detectable levels of HCV RNA in clinical patients but not necessarily in blood donors.     

Prevalence of anti-HCV antibody in family members of anti-HCV-positive patients with acute and chronic liver disease.

Presence of circulating anti-hepatitis C antibody (anti-HCV) was screened in 201 Thai patients with acute and chronic liver disease who presented to Ramathibodi and Phya Thai Hospitals during 1984-1990. Of these, 29 patients (14.4%) were positive for anti-HCV. Circulating anti-HCV was determined in 92 family members (20 spouses, 72 household contacts) of these index cases and was detected in 5 contacts (2 spouses, 2 daughters and 1 mother) of 3 index cases. The overall prevalence of anti-HCV among the contacts was 5.4% (5/92) and it was higher in sexual partners (2/20, 10.0%) compared to other household contacts (3/72, 4.2%) but this was not statistically significant (p = 0.297). The anti-HCV-positive contacts were significantly older (mean +/- SD = 61.4 +/- 14.4) than the other contacts either comparing within the same families (26 +/- 16.5; p = 0.012) or all studied families (25.1 +/- 13.3; p = 0.006). One anti-HCV-positive contact had hepatocellular carcinoma, one had unexplained elevation of serum aminotransferase and the remaining 3 had no clinical or laboratory evidence of liver disease. All of the 3 index cases with anti-HCV-positive contacts, had chronic liver disease (2 cirrhosis, 1 chronic persistent hepatitis) and the prevalence of anti-HCV in these families (8/13, 61.5%) was significantly higher than the remaining 26 families (26/108, 24.1%) (p = 0.008). The results of this study suggest that sexual and other intrafamilial personal contact may be important for HCV transmission. Duration of close contact and family relationships appear to determine this mode of HCV transmission.     

MEIA法与ELISA法检测丙型肝炎病毒抗体结果比较

目的比较MEIA法与ELISA法检NHCV抗体的结果,以及2种方法在实际应用中的区别。方法收集152例ELISA法检NHCV抗体阳性的血清标本,并采用MEIA法检测,对2种检测方法的结果进行比较。结果152例ELISA法检NHCV抗体阳性的血清标本采用MEIA法检测后,144例为阳性,8例为阴性。结论ELISA法操作简单、灵敏度高,但在实际操作中,ELISA检测HCV抗体结果影响因素较多,易出现假阳性问题;MEIA法检测HCV抗体较ELISA法准确、灵敏、快速。     

Gold immunoblot analysis of IgM-specific antibody in the diagnosis of human leptospirosis.

An immunoblot for the detection of leptospirosis was developed in our laboratory. Antigen prepared from Leptospira interrogans serovar bataviae was dotted onto nitrocellulose paper and blocked with skim milk. Test and control sera diluted 1:20 were applied to the dot, incubated, and washed. Anti-human IgM colloidal gold conjugate was added and the dots were washed. A positive reaction was shown by the development of a pink dot against a white background. The test was performed on 62 sera that tested positive for leptospirosis by a microagglutination (MA) test, on 40 sera that were positive by an indirect hemagglutination (IHA) test, and on sera from forty healthy blood donors. Four sera from the blood donors showed a faint pink dot, but the remainder showed a colorless reaction. All 62 sera that tested positive by MA were positive by this new test, while 95% of the 20 sera that tested positive by IHA were positive. Tests for IgG antibody were performed on 20 sera positive by MA using protein A-colloidal gold conjugate, and all showed weak reactivity. The results confirmed previous findings that most antibodies present in leptospirosis patients are of the IgM type. The ELISA takes three hours to perform, but the gold immunoblot can be completed in 30 min. In addition, the test blot can be kept as a permanent record, and is a significant improvement over existing tests.     

A study of antigen-specific anti-cytomegalovirus antibody reactivity in patients with systemic sclerosis and concomitant anti-Ro52 antibodies

Rheumatology International - Anti-Ro52 autoantibody (autoAb), highly prevalent in Sjogren’s syndrome (SjS) and systemic lupus erythematosus (SLE), is also frequent in systemic sclerosis...     

Detection of anti-HAY antibody with dot immunogold filtration assay

AIM: To establish a rapid, sensitive and specific immunogold assay for detection of hepatitis A virus infection.METHODS: Rabbit monoclonal antibodies to anti-human IgM and IgG (Dako) were dotted on a nitrocellulose membrane (NCM) respectively to capture the human sera IgM and IgG. Then the captured antibodies would conjugate to HAV antigen, which was revealed by mouse anti-HAV IgG conjugated to gold particles. Final results were assessed by blind method.RESULTS: Sera from 96 patients with acute hepatitis were used for our study. Compared with well-recognized standard (Abbott Laboratory, USA), the sensitivity and specificity of IgM-DIGFA (self-made) were 91.3 % (42/46) and 96.0 %(48/50), and those of IgM-ELISA (Kehua, Shanghai) were 97.8 % (45/46) and 100.0 % (50/50). The identical results were produced from the study with reagents at different conditions, and the study was repeated in 15 negative sera and 10 positive sera. The serum anti-HAV IgG was tested with DIGFA at the same time. In comparison with ELISA,the sensitivity and specificity of DIGFA for IgG anti-HAV were 87.2 % (41/47) and 91.8 % (45/49), respectively.CONCLUSION: This assay can detect anti-HAV IgM and IgG simultaneously, and be done within 3 minutes. The simplicity, rapidity and specificity of the assay were useful for screening and epidemiological study.