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Objective: There are three major serologic markers for hepatitis B virus (HBV) infection: hepatitis B surface antigen (HBsAg); hepatitis B surface antibody (anti-HBs); and hepatitis B core antibody (anti-HBc). HBV screening programs, however, often test only HBsAg and anti-HBs, missing those individuals who have anti-HBc as the only detectable marker. Isolated anti-HBc can represent chronic infection in which HBsAg is not detectable by serology. We, therefore, investigated the prevalence of isolated anti-HBc in an ethnic community at moderate to high risk for HBV infection., Results: Of 7157 Korean American adults in New Jersey, 2736 (38.2%) lacked anti-HBs, potentially susceptible to HBV. Of these 2736 subjects, 771 subjects had anti-HBc. The prevalence of isolated anti-HBc increased with age: 0.8% (age 21-30); 2.4% (age 31-40); 6.05% (age 41-50); 11.7% (age 51-60); 18.3% (age 61-70); and 24.5% (age 71-91). Similarly, the percentage of the individuals with isolated anti-HBc in anti-HBs lacking subjects showed a striking age dependence. We conclude that serologic HBV screening should include anti-HBc to accurately assess the prevalence of HBV exposure. Serologic screening with only HBsAg and anti-HBs may overestimate the prevalence of non-immune population. It can also underestimate the prevalence of HBV and increase the risk of HBV reactivation during immunosuppression.

Background: Previous contact with Hepatitis B virus (HBV) is common in patients undergoing hemodialysis. Literature has shown conflicting results on the risk of HBV reactivation in kidney transplant (KT) recipients with serologic evidence of past HBV infection., Methods: We reviewed 631 consecutive KT recipients and selected 70 patients simultaneously HBsAg negative and anti-HBc positive before KT, regardless of hepatitis B surface antibody (anti-HBs) status. Demographic characteristics, coinfection with other viruses, the presence of a previous KT, induction and maintenance immunosuppression, length of follow up, biopsy-proven acute rejection episodes, incidence of impaired liver function, and causes of graft loss and mortality were collected. Hepatitis B virus reactivation was defined as detection of HBV DNA viral load >2000 IU/mL during follow up. Outcome data included HBV reactivation episodes, graft function, and patient survival., Results: Median follow-up was 151 months; 91.4% of patients were positive to anti-HBs prior to KT. No patient received HBV prophylaxis and 11 patients (15.7%) received rituximab as part of induction therapy. Anti-HBs titers remained stable in all patients throughout the observation period but two patient showed evidence of HBV reactivation after KT., Conclusion: Hepatitis B virus reactivation in HBsAg-negative and anti-HBc-positive after KT is rare but possible. We suggest evaluating HBV serologies, HBV DNA viral load, and liver enzymes before KT and routinely monitoring serologic HBV markers after KT. As only two patients experienced HBV reactivation, it is neither possible to define risk factors for HBV reactivation nor to evaluate the impact of different immunosuppressants or the benefit of prophylactic regimens. Further studies regarding HBV reactivation in solid organ transplant recipients are necessary., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Given the shortage of suitable liver grafts for liver transplantation, proper use of hepatitis B core antibody-positive livers might be a possible way to enlarge the donor pool and to save patients with end-stage liver diseases. However, the safety of hepatitis B virus core antibody positive (HBcAb + ) donors has been controversial. Initial studies were mainly conducted overseas with relatively small numbers of HBcAb + liver recipients, and there are few relevant reports in the population of mainland China. We hypothesized that the safety of HBcAb + liver grafts is not suboptimal., Aim: To evaluate the safety of using hepatitis B virus (HBV) core antibody-positive donors for liver transplantation in Chinese patients., Methods: We conducted a retrospective study enrolling 1071 patients who underwent liver transplantation consecutively from 2005 to 2016 at West China Hospital Liver Transplantation Center. Given the imbalance in several baseline variables, propensity score matching was used, and the outcomes of all recipients were reviewed in this study., Results: In the whole population, 230 patients received HBcAb + and 841 patients received HBcAb negative (HBcAb - ) liver grafts. The 1-, 3- and 5-year survival rates in patients and grafts between the two groups were similar (patient survival: 85.8% vs 87.2%, 77.4% vs 81.1%, 72.4% vs 76.7%, log-rank test, P = 0.16; graft survival: 83.2% vs 83.6%, 73.8% vs 75.9%, 70.8% vs 74.4%, log-rank test, P = 0.19). After propensity score matching, 210 pairs of patients were generated. The corresponding 1-, 3- and 5-year patient and graft survival rates showed no significant differences. Further studies illustrated that the post-transplant major complication rates and liver function recovery after surgery were also similar. In addition, multivariate regression analysis in the original cohort and propensity score-matched Cox analysis demonstrated that receiving HBcAb + liver grafts was not a significant risk factor for long-term survival. These findings were consistent in both HBV surface antigen-positive (HBsAg + ) and HBsAg negative (HBsAg - ) patients.Newly diagnosed HBV infection had a relatively higher incidence in HBsAg - patients with HBcAb + liver grafts (13.23%), in which HBV naive recipients suffered most (31.82%), although this difference did not affect patient and graft survival ( P = 0.50 and P = 0.49, respectively). Recipients with a high HBV surface antibody (anti-HBs) titer (more than 100 IU/L) before transplantation and antiviral prophylaxis with nucleos(t)ide antiviral agents post-operation, such as nucleos(t)ide antiviral agents, had lower de novo HBV infection risks., Conclusion: HBcAb + liver grafts do not affect the long-term outcome of the recipients. Combined with proper postoperative antiviral prophylaxis, utilization of HBcAb + grafts is rational and feasible., Competing Interests: Conflict-of-interest statement: All authors declare no conflicts of interest related to this article.

The hepatitis B virus (HBV) envelope contains small (S), middle (M), and large (L) proteins. PreS1 of the L protein contains a receptor-binding motif crucial for HBV infection. This motif is highly conserved among 10 HBV genotypes (A-J), making it a potential target for the prevention of HBV infection. In this study, we successfully generated a neutralizing human monoclonal antibody (mAb), 1A8 (IgG1), that recognizes the receptor-binding motif of preS1 using a phage-displayed human synthetic Fab library. Analysis of the antigen-binding activity of 1A8 for different genotypes indicated that it can specifically bind to the preS1 of major HBV genotypes (A-D). Based on Bio-Layer interferometry, the affinity ( K D ) of 1A8 for the preS1 of genotype C was 3.55 nM. 1A8 immunoprecipitated the hepatitis B virions of genotypes C and D. In an in vitro neutralization assay using HepG2 cells overexpressing the cellular receptor sodium taurocholate cotransporting polypeptide, 1A8 effectively neutralized HBV infection with genotype D. Taken together, the results suggest that 1A8 may neutralize the four HBV genotypes. Considering that genotypes A-D are most prevalent, 1A8 may be a neutralizing human mAb with promising potential in the prevention and treatment of HBV infection.

Background and Study Aims: Occult hepatitis B infection (OBI) is known to be mostly prevalent in chronic hepatitis C (CHC) patients and OBI reactivation might be life-threatening in patients undergoing interferon (IFN)-free direct acting antiviral (DAA) therapy. As previous studies have revealed a relationship between OBI and non-response to IFN-based antiviral therapy, the aim of the current study was to determine if there was a higher prevalence of OBI in IFN non-responders than responders., Patients and Methods: This retrospective cross-sectional study was conducted in CHC patients who had previously received IFN-based antiviral therapy. Serum samples of 100 HBsAg negative CHC patients were tested for HBV DNA, anti-HBc IgG, anti-HBs, ALT and AST. The presence of OBI was compared between 50 IFN responders and 50 IFN non-responders. Patients with a history of previous HBV infection, patients with evidence of cirrhosis and patients who had received IFN therapy within the last one year were excluded from the study., Results: Anti-HBc IgG positivity was determined in 53% of the patients. HBV DNA positivity, indicating OBI was determined in 1 (1%) patient. This patient was anti-HBc IgG positive, anti-HBs negative, ALT and AST levels were normal. The HBV DNA and anti-HBc IgG positivity rates were higher in the non-responder group than in the responder group, but the difference was not statistically significant (p = 0.31 and p = 0.07 respectively)., Conclusion: According to the results of this study, the prevalence of OBI is lower than expected amongst CHC patients in Turkey and it may not be necessary to apply routine screening to IFN non-responders for OBI infection before DAA therapy. However, there is a need for multicentre studies with larger patient series., (Copyright © 2018 Pan-Arab Association of Gastroenterology. Published by Elsevier B.V. All rights reserved.)

Circulating immune complexes (CICs) are produced during the immune response. It is more clinically important to establish a general and efficient CICs dissociation technique for the detection of antigens for CICs other than the detection of free antigens in the serum. Polyethylene glycol (PEG) two-precipitation separation and glycine-HCl as a buffer system were employed to develop a general and efficient buffer dissociation technique to separate CICs from serum and dissociate antigens from CICs. The measurement value of new PEG two-precipitation separation technique was higher than traditional PEG precipitation separation technique. There were slight differences in the dissociation conditions of HCV Core-IC, HIV P24-IC, Ins-IC and TG-IC as compared to HBsAg-IC. The detection of antigens in HBsAg-IC, HCV Core-IC, HIV P24-IC, Ins-IC and TG-IC with this technique was superior to that with HCl Dissociation, Trypsin Digestion or Immune Complex Transfer technique. PEG two-precipitation dissociation technique may reduce macromolecular protein and the adhesion of free antigens during the co-precipitation, which increases the efficiency of separation and precipitation of CICs. This technique also avoids the damage of reagents to antigens, assuring the repeatability, reliability and validity. Thus, this technique is application in samples negative or positive for free antigens., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Hepatitis B virus (HBV) reactivation has been observed following interferon (IFN)-based treatment in HBV/hepatitis C virus (HCV) co-infected patients. Recent reports suggest that reactivation may also occur in both hepatitis B surface antigen (HBsAg)-positive and HBsAg-negative patients during HCV treatment with direct-acting antivirals (DAAs)., Aim: To investigate the rate of patients with HBV reactivation during IFN-based and IFN-free HCV treatment in a large real-world cohort., Methods: A total of 848 patients with chronic hepatitis C were treated with different combinations of DAAs. Among patients with available outcome and HBV data, there were 272 patients hepatitis B core antibody (HBcAb)-positive (HBsAg-positive, n=9; HBsAg-negative, n=263), and 536 were HBcAb-negative. All HBcAb-positive patients were tested for HBV DNA at the end of DAA therapy and alanine transaminase (ALT) levels were frequently measured during therapy and follow-up., Results: Seventy-three percent (n=192/263) of HBsAg-negative/HBcAb-positive patients had elevated ALT levels at baseline, which declined to normal values in all but 18 patients, and no HBV reactivation was observed. Eight patients had detectable but not quantifiable HBV DNA (<20 IU/mL) at end of treatment, but none were associated with elevated ALT. Five of nine HBsAg-positive/HBcAb-positive patients experienced transient or permanent HBV reactivation, three of whom required nucleos(t)ide treatment during (n=1) or after (n=2) DAA therapy., Conclusions: HBV reactivation was not observed in HBsAg-negative/HBcAb-positive patients but common in HBsAg-positive/HBcAb-positive patients treated with different combinations of DAAs for HCV., (© 2017 John Wiley & Sons Ltd.)

Vaccination is the most effective and well-tolerated method of conferring long-term protection against hepatitis A and B viruses (HAV; HBV). Long-term studies are required to characterize the duration of protection and need for boosters. Following primary immunization of 150 and 157 healthy adults with 3-doses of combined hepatitis A/hepatitis B vaccine (HAB; Twinrix™, GSK Vaccines, Belgium) at 0-1-6 months in 2 separate studies, we measured vaccine-induced antibody persistence against HAV and HBV annually for 20 y (Study A: NCT01000324; Study B: NCT01037114). Subjects with circulating anti-HAV antibodies < 15 mIU/mL or with anti-hepatitis B surface antigen < 10 mIU/mL were offered an additional monovalent hepatitis A and/or B vaccine dose (Havrix™/Engerix™-B, GSK Vaccines, Belgium). Applying the immunogenicity results from these studies, mathematical modeling predicted long-term persistence. After 20 y, 18 and 25 subjects in studies A and B, respectively, comprised the long-term according-to-protocol cohort for immunogenicity; 100% and 96.0% retained anti-HAV antibodies ≥ 15 mIU/mL, respectively; 94.4% and 92.0% had anti-HBs antibodies ≥ 10 mIU/mL, respectively. Between Years 16-20, 4 subjects who received a challenge dose of monovalent hepatitis A vaccine (N = 2) or hepatitis B vaccine (N = 2), all mounted a strong anamnestic response suggestive of immune memory despite low antibody levels. Mathematical modeling predicts that 40 y after vaccination ≥ 97% vaccinees will maintain anti-HAV ≥ 15 mIU/mL and ≥ 50% vaccinees will retain anti-HBs ≥ 10 mIU/mL. Immunogenicity data confirm that primary immunization with 3-doses of HAB induces persisting anti-HAV and anti-HBs specific antibodies in most adults for up to 20 y; mathematical modeling predicts even longer-term protection.

Background: Although an isolated anti-hepatitis B virus (HBV) core antibody (anti-HBc) serological profile is frequent in human immunodeficiency virus (HIV)-infected patients, data on HBV vaccination in these patients are scarce., Methods: A prospective multicenter study was conducted to assess the immunogenicity of HBV vaccination in 54 patients with an isolated anti-HBc profile and undetectable HIV load. They were vaccinated with 1 dose (20 µg) of recombinant HBV vaccine. Those with an anti-HBV surface antibody (anti-HBs) level of <10 mIU/mL 4 weeks after vaccination received 3 additional double doses (40 µg) at weeks 5, 9, and 24., Results: At week 4, 25 patients (46%) were responders. Only the ratio of CD4(+) T cells to CD8(+) T cells was associated with this response in multivariate analysis (odds ratio for +0.1, 1.32; 95% confidence interval, 1.07-1.63; P = .008). At week 28 and month 18, 58% of these patients (14 of 24) and 50% (10 of 20), respectively, maintained anti-HBs level of ≥10 mIU/mL.Among nonresponding patients at week 4, who received further vaccinations, 89% (24 of 27) and 81% (21 of 26) had an anti-HBs level of ≥10 mIU/mL at week 28 and month 18, respectively. The preS2-specific interferon γ T-cell response increased between week 0 and week 28 in patients who finally responded to reinforced vaccination (P = .03)., Conclusions: All of the patients with an isolated anti-HBc profile who did not have an anti-HBs titer of >100 mIU/mL 4 weeks after a single recall dose of HBV vaccine should be further vaccinated with a reinforced triple double-dose scheme., (© The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.)

Objective: To examine the immune memory status to hepatitis B vaccine among infants with normal or high antibody response to primary vaccination, 5 years after the primary vaccination and the risk factors associated with the immune memory., Methods: Titers of the antibody against hepatitis B surface antigen (anti-HBs) were detected, five years after the primary vaccination among children who appeared normal or high response to hepatitis B primary vaccination in infancy. Those whose anti-HBs titers were low than protective level (10 mIU/ml) were given a challenge dose of hepatitis B vaccine and titers of anti-HBs were detected 14 days after the challenge. Positive rate and geometric mean titer (GMT) of anti-HBs were calculated. Level of the anti-HBs titers after primary vaccination, at following-up and after the challenge periods were divided into different levels, respectively. Risk factors associated with the levels of anti-HBs titer after the challenge were examined by univariate analysis that and multivariable analysis., Results: Anti-HBs waned to the level below protective standard among 37.98% of the children with normal or high antibody response to hepatitis B primary vaccination; among those children whose anti-HBs were below the protection standard. The seroconversion rate and GMT of anti-HBs after the challenge dose were 98.95% (757/765) and 2 811.69 mIU/ml [95% Confidence Interval (CI) :2 513.55-3 145.19 mIU/ml] , respectively. Titers and levels of anti-HBs after the challenge, appeared an increase with anti-HBs after primary vaccination and the anti-HBs in the following-up (F=5.46, 10.23 respectively; P<0.000 1 for both) periods. Results from the multivariable analysis showed that gender, premature birth and birth weight were factors insignificantly associated with the anti-HBs titers after the dose of challenge, while the anti-HBs levels were independently associated with the levels of anti-HBs titer after the challenge [OR = 1.001 (95%CI: 1.000-1.002) , P<0.001; OR=1.28 (95%CI: 1.81-1.39) , P<0.001]at the following-up periods., Conclusions: Strong immune memory could be found among those children with normal or high responses to hepatitis B vaccination, 5 years after the primary vaccination. The intensity of immune memory might be associated with the anti-HBs titer after primary vaccination as well as the anti-HBs titers before the challenge dose was given.

Background: At present, the scarcity of organs is one of the major problems in transplantation. We present our own experience in expanding the acceptance criteria for deceased donors. We aimed to evaluate the potential impact of using hepatitis B virus (HBV) core antibody (anti-HBc)-positive donor grafts on the long-term liver transplantation results., Material and Methods: We retrospectively analyzed data from 324 consecutive liver transplantation procedures performed in the largest transplant center in Poland, between January 2007 and February 2012. These patients included 36 who had received a liver from anti-HBc-positive donors., Results: The presence of anti-HBc antibodies in the donor did not have a significant negative impact on the overall 3-year patient or graft survival rates. Among recipients with pre-transplant HBV infection, patients who received an organ from an anti-HBc-positive donor tended to have an improved survival rate compared with patients who received liver transplants from seronegative donors (p=0.080). However, there was a statistically significant reduction in graft survival (p=0.035) in the recipients without serological markers of HBV infection who received liver transplants harvested from anti-HBc-positive donors., Conclusions: These results confirm the validity of extending the acceptance criteria to anti-HBc-positive donors, particularly for patients with a prior HBV infection. Despite the increased risk of graft loss, liver transplantation from anti-HBc-positive donors among recipients not infected with HBV may also be considered in appropriate cases.

A biosensor is an analytical device used for the detection of analytes, which combines a biological component with a physicochemical detector. Recently, an increasing number of biosensors have been used in clinical research, for example, the blood glucose biosensor. This review focuses on the current state of biosensor research with respect to efficient, specific and rapid detection of hepatitis B virus (HBV). The biosensors developed based on different techniques, including optical methods (e.g., surface plasmon resonance), acoustic wave technologies (e.g., quartz crystal microbalance), electrochemistry (amperometry, voltammetry and impedance) and novel nanotechnology, are also discussed.

Aim: Determine the frequency of antibodies to surface (anti-HBs) and core (anti-HBc) antigens of hepatitis B in population of St. Petersburg of various age for evaluation of protective herd immunity against hepatitis B virus (HB)., Materials and Methods: Blood sera of 970 individuals (491 males, 479 females) of 10 age groups from 0 to 50 years and older were examined for the presence of anti-HBs and anti-HBc IgG by enzyme immunoassay using commercial diagnostic test-systems., Results: In general anti-HBs at the level of 5 mIU/ml and above were detected in 603 of the examined individuals (62.2%). Anti-HBs at the level of 10 mIU/ml and above were detected in 53.9%. The frequency of anti-HBs in protective titers in males and females in general turned out to be similar (52.6% and 55.2%, respectively). Juxtaposition of age-specific parameters of seroprotection and acute HB morbidity in St. Petersburg revealed an inverse correlation of medium strength (r = - 0.54)., Conclusion: Results of the study confirm high effectiveness of the program of HB vaccine prophylaxis in St. Petersburg and emphasize the necessity of further implementation of broad measures of population immunization against HB.

Enzyme immunoassay was used to determine the presence of immunoglobulins class G to Toxocara canis antigens in the sera of children and adolescents (hereinafter referred to as children) with allergic and bronchopulmonary diseases from HIV infection and hepatitis B and C risk groups. A total of 422 dwellers of the Republic of Altai, including 144 subjects aged 1 to 17 years, were examined. Toxocara antibodies were found in 18.8 +/- 3.3% of the children and in 21.9 +/- 2.5% of the adults. The infection rate in children with bronchopulmonary and allergic diseases was 27.1 +/- 5.8 and 14.3 +/- 5.0%, respectively; that in the hepatitis B and C risk groups was 13.1 +/- 6.2%. The children (n = 6) from the HIV infection risk group were seronegative. The infection rate in the adults from the HIV infection and hepatitis risk group was 19.2 +/- 3.5 and 24.3 +/- 3.5%, respectively. Diagnostic antibody titers in the children and adults were determined in 9.0 +/- 2.3 and 8.3 +/- 1.6%, respectively. Immunological assays should be used to rule out toxocariasis in the examinees. If there are seropositive results, specific antiparasitic threatment should be performed.

In the present study, we have focused our attention to prepare molecular imprinted composite cryogel membranes for purification of hepatitis B surface antibody (anti-HBs) by fast protein liquid chromatography. Before the preparation of the molecular imprinted composite cryogel membranes (MI-CMs) by free radical polymerization at sub-zero temperature, we have synthesized and characterized the anti-HBs imprinted particles. Then, the cryogel membranes (CMs) were characterized by swelling test, scanning electron microscopy and Fourier transform infrared spectroscopy. Prior to chromatographic purification studies, the effective parameters on the anti-HBs adsorption process were evaluated by investigating the dependency of the adsorption capacity on flow-rate, anti-HBs concentration, contact time and ionic strength. The maximum anti-HBs adsorption capacity was calculated as 701.4 mIU/g CM. The selectivity of the MI-CMs was shown by competitive adsorption of anti-HBs, total anti-hepatitis A antibody (anti-HAV) and total immunoglobulin E (IgE) adsorption studies. The MI-CMs have relative selectivity coefficients as 5.45 for anti-HBs/total anti-HAV and 9.05 for anti-HBs/total IgE, respectively. The phosphate buffer solution (pH 7.4) containing 1.0M NaCl was used for elution, almost completely, of adsorbed anti-HBs molecules. The MI-CMs could be used many times without any significant decrease in the adsorption capacity. The chromatographic purification performances of the MI-CMs were also investigated. The chromatographic parameters such as capacity and separation factors, the theoretical plate number and resolution of the MI-CMs were calculated as 5.48, 6.02, 1153.9, and 1.72 for anti-HBs molecules, respectively. As a conclusion, we can say that the MI-CMs could be used for specific purification of anti-HBs from anti-HBs positive human plasma., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Cytotoxic T lymphocyte antigen 4 (CTLA-4) is an inhibitory T cell receptor predominately expressed on activated T cells and plays an important role in regulation of specific T cell responses to viral infection. The woodchuck model is an informative animal model for hepatitis B virus (HBV) infection. In this study, the extracellular region of woodchuck CTLA-4 (wCTLA-4) was cloned and the fusion protein of GST-wCTLA-4 was expressed and purified. Polyclonal antibody against GST-wCTLA-4 (anti-GST-wCTLA-4) was prepared. The full length wCTLA-4 protein expressed in transfected baby hamster kidney cells was detected by anti-GST-wCTLA-4 in western blot analysis and immunofluorescence staining. Anti-GST-wCTLA-4 provides a useful tool to study the role of CTLA-4 in T-cell response in the woodchuck model. Further, the blocking of CTLA-4 with anti-GST-wCTLA-4, as a novel therapy approach for chronic hepatitis B virus infection, could be studied in woodchuck model now., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Objective: To develop an amplified luminescent proximity homogeneous immunoassay (AlphaLISA) kit for the detection of human hepatitis B virus e antibody (HBeAb)., Methods: The neutralizing and competitive inhibition method was used to develop the AlphaLISA kit for detection of serum HBeAb., Results: The working range of the kit was 0.003-16 NCU/ml with a sensitivity up to 0.003 NCU/ml. The intra- and inter-assay coefficient of variation was 5.3% and 6.8%, respectively. The kit showed no cross-reaction with HBcAb, and comparison of the detection results with those of a commercially available Elecsys HBeAb kit (Roche) for 136 samples showed a correlation coefficient of 0.961., Conclusion: The AlphaLISA kit for HBeAb detection meets the clinical requirements for detection HBeAb in human serum.

Quantification of hepatitis B surface antigen (HBsAg) or relative in vitro potency in the final vaccines is a prerequisite for hepatitis B vaccine batch release. The commercial kit for automated analysis (AxSYM) is expensive, and an alternative is required for the estimation of HBsAg in hepatitis B vaccines. Mouse monoclonal antibodies (MAbs) specific for HBsAg were developed and characterized. One of the monoclonal antibodies (HBs06) was used in development of an immunocapture ELISA (IC-ELISA) as an unlabeled capture antibody and biotin-labeled detection antibody. The IC-ELISA was standardized and validated using experimental hepatitis B vaccine batches with various HBsAg concentrations per dose and commercial vaccines. The vaccine was treated with an alkaline solubilizer to desorb the HBsAg from Algel-adjuvanted vaccines before testing, and the sensitivity of the test was 5 ng/ml. A good correlation could be observed between the HBsAg estimates derived by both formats, except for the higher HBsAg concentration range, where the IC-ELISA format could estimate closer to the actual values than AxSYM. There was a significant correlation between the estimated relative potencies of the two methods. There was lack of correlation between the in vivo potency and the relative in vitro potency. However, the estimates of IC-ELISA were comparable to the in vivo values when compared with the estimates of AxSYM. The IC-ELISA can therefore be considered to be a reliable test for deriving in vitro relative potency and antigen concentration in vaccine batches for batch control and release.

In Canada, universal and publicly funded hepatitis B immunization programs have been available since 1998 in all provinces and territories. This present study estimates the proportion of having vaccine-induced immunity to hepatitis B virus (HBV) infection and its associated determinants among street-involved youth aged at 15-24 years old in Canada using the data collected by the Enhanced Surveillance of Canadian Street Youth. Vaccine-induced immunity was identified by blood test results of anti-HBc negative and anti-HBs positive. Of the 4,035 participants included in this study, the overall proportion of those with vaccine-induced immunity to HBV was 51.7% during the study period compared to over 90% among the general adolescent population. The proportion of street-involved youth immunized with HBV vaccine increased from 34.7% in 1999 to 64.4% in 2005. Immunity was higher among females (aOR = 1.43, 1.17-1.75) and among those with a reported history of sexually transmitted infection (aOR = 1.30, 1.03-1.63). The proportion of youth with the immunity decreased as age increased (aOR = 0.78, 0.76-0.81, per year increase). Despite an overall increase in the proportion of Canadian street-involved youth with vaccine-induced immunity to HBV, the proportion was still significantly lower than that observed in the general adolescent population. This highlights the need to improve the access to basic health care and the immunization programs to HBV for street-involved youth through creative outreach programs and other multi-faceted approaches.

Hepatitis B virus (HBV) infections represent a global health problem, since these account for 350 million chronic infections worldwide that result in 500,000-700,000 deaths each year. Control of viral replication and HBV-related disease and mortality are of utmost importance. Because the currently available antiviral therapies all have major limitations, new strategies to treat chronic HBV infection are eagerly awaited. Six single-domain antibodies (VHHs) targeting the core antigen of HBV (HBcAg) have been generated and three of these bound strongly to HBcAg of both subtype ayw and adw. These three VHHs were studied as intrabodies directed towards the nucleus or the cytoplasm of a hepatoma cell line that was co-transfected with HBV. A speckled staining of HBcAg was observed in the cytoplasm of cells transfected with nucleotropic VHH intrabodies. Moreover, an increased intracellular accumulation of hepatitis B e antigen (HBeAg) and a complete disappearance of intracellular HBcAg signal were observed with nuclear targeted HBcAg-specific VHHs. These results suggest that HBcAg-specific VHHs targeted to the nucleus affect HBcAg and HBeAg expression and trafficking in HBV-transfected hepatocytes.

Hepatitis B surface antibody imprinted poly(hydroxyethyl methacrylate-N-methacryloyl-L-tyrosine methyl ester) particles were prepared for the purification of hepatitis B surface antibody from human plasma. N-methacryloyl-L-tyrosine methyl ester was chosen as a complexing agent for hepatitis B surface antibodies. Hepatitis B surface antibody imprinted poly(hydroxyethyl methacrylate-N-methacryloyl-L-tyrosine methyl ester) particles were characterized by surface area measurements, swelling test, scanning electron microscopy, elemental analysis, and Fourier transform infrared spectroscopy. Ethylene glycol (1.0M) was used as desorption agent. Adsorption studies were performed from hepatitis B surface antibody and anti-hepatitis A antibody positive human plasma. Effects of antibody concentration, contact time, N-methacryloyl-L-tyrosine methyl ester content and temperature on the adsorption capacity were investigated. The amount of hepatitis B surface antibody adsorbed per unit mass increased with increasing hepatitis B surface antibody concentration, then reached saturation. Maximum hepatitis B surface antibody adsorption amount was 21.4 mIU/mg. Adsorption process reached the equilibrium in 60 min. Competitive adsorption of hepatitis B surface antibody, total anti-hepatitis A antibody and total immunoglobulin E was investigated for showing the selectivity. Hepatitis B surface antibody-imprinted particles could adsorb hepatitis B surface antibody 18.3 times more than anti-hepatitis A antibody and 2.2 times more than immunoglobulin E. It can be concluded that hepatitis B surface antibody-imprinted particles have significant selectivity for hepatitis B surface antibody.

Expenditure on screening blood donations in developing countries can be reduced by testing donations in pools. This study evaluated serological screening in pools for hepatitis B virus (HBV) at the Israeli national blood bank and a hospital blood bank in Gaza, the Palestinian Authority. The accuracy of HBV surface antigen (HBsAg) enzyme immunoassay performed on pools of 3-24 samples was compared with individual tests. Delay in detecting positive samples due to dilution in pools and the possibility of antibody-antigen neutralization were analyzed. The sensitivity of pooled testing for HBsAg was 93-99%, prolonging the window period by 5 days (8.3%). Neutralization of HBsAg by hepatitis B surface antibodies (anti-HBs) could be minimized by testing immediately after pooling. Serological testing for HBsAg in pools may be performed using manually created pools of up to six samples, with 5% loss in sensitivity and a risk of neutralization by anti-HBs present in the donor population. Pooling can therefore be considered as an option only in countries with a low prevalence of HBV.

Aim: To construct a prokaryotic plasmid expressing truncated duck hepatitis B virus core protein (DHBc(1-214)), purify the recombinant protein, and to develop polyclonal antibodies against DHBc., Methods: DHBc(1-214) was cloned into vector pRSET-B, then expressed in E.coli Rosetta(DE3) pLacI induced by IPTG. The recombinant protein was purified using Ni-NTA spin column. Polyclonal antibody was developed by immunizing BALB/c mice with the purified recombinant protein, and their sensitivity and specificity were tested using enzyme-linked immunosorbent assay, immunohistochemical staining and Western blot analysis., Results: Recombinant plasmid expressing truncated DHBc(1-214) was successfully constructed. A protein of 28,000 was expressed and purified. Polyclonal serum antibody with a high specificity was obtained by immunizing BALB/c mice with the purified recombinant protein., Conclusion: The truncated recombinant DHBc(1-214) developed in this study is purified and shown strong antigenecity. The polyclonal antibody against DHBc protein is generated by regular immunization method, demonstrating both high sensitivity and specificity. The protein and the antibody can be used for further clinical examination and research of DHBV.

Objectives: We measured the prevalence of hepatitis B virus (HBV) immunization and HBV infection among men aged 23 to 29 years who have sex with men., Methods: We analyzed data from 2834 men who have sex with men in 6 US metropolitan areas. Participants were interviewed and tested for serologic markers of immunization and HBV infection in 1998 through 2000., Results: Immunization prevalence was 17.2%; coverage was 21.0% among participants with private physicians or health maintenance organizations and 12.6% among those with no source of health care. Overall, 20.6% had markers of HBV infection, ranging from 13.7% among the youngest to 31.0% among the oldest participants. Among those susceptible to HBV, 93.5% had regular sources of health care, had been tested for HIV, or had been treated for a sexually transmitted disease., Conclusions: Although many young men who have sex with men have access to health care, most are not immunized against HBV. To reduce morbidity from HBV in this population, providers of health care, including sexually transmitted disease and HIV prevention services, should provide vaccinations or referrals for vaccination.

Antibodies have been one of the proteins widely expressed in tobacco plants for pharmaceutical purposes, which demand contaminant free preparations. Rubisco constitutes 40-60% of tobacco leaf soluble proteins; therefore it is the major potential protein contaminant of plantibodies, while mycotoxins are toxic compounds that could be introduced during the biomass production and post-harvest stages with important consequences to human health. The objective of this paper was to investigate whether Rubisco and mycotoxins are present in Plantibody HB-01 preparations used in the immunopurification of the hepatitis B surface antigen. Rubisco was purified from Nicotiana tabacum yielding 154 microg of protein per gram of leaves and purity over 95%. Among mouse monoclonal antibodies generated against this enzyme, the CBSS.Rub-2 was selected for its immunodetection. It recognizes a conserved sequential epitope of Rubisco large subunit with an affinity constant of 0.13 x 10(8)M(-1). Rubisco quantification limit was 1 microg spreading to the measurement of this contaminant less than 4% of plantibodies samples. Additionally, according to a Reverse Phase-HPLC used to measure the level of adventitiously introduced contaminants, it can be concluded that aflatoxins B1, B2, G1 and G2 were undetected in the purified Plantibody HB-01 samples.

Aim: To evaluate the prevalence of isolated anti-HBc in patients with chronic hepatitis C virus (HCV) infection, and its relation to disease severity., Methods: We screened all patients with chronic HCV infection referred to King Faisal Specialist Hospital and Research Center for hepatitis B surface antigen (HBsAg), antibody to hepatitis B surface antigen (anti-HBs), and anti-HBc. One hundred and sixty nine patients who tested negative for both HBsAg and anti-HBs were included in this study., Results: Pathologically, 59 had biopsy-proven cirrhosis and 110 had chronic active hepatitis (CAH). Of these 169 patients, 85 (50.3%) tested positive for anti-HBc. Patients with CAH had significantly higher prevalence of isolated anti-HBc than patients with cirrhosis, 71 (64.5%) and 14 (23.7%) respectively (P < 0.001). Twenty-five patients were tested for HBV DNA by qualitative PCR. The test was positive in 3 of them (12%; occult HBV infection)., Conclusion: Isolated anti-HBc alone is common in Saudi patients with chronic HCV infection, and is significantly more common in those with CAH than those with cirrhosis. Therefore, a screening strategy that only tests for HBsAg and anti-HBs in these patients will miss a large number of individuals with isolated anti-HBc, who may be potentially infectious.

Objective: To optimize the adsorption condition of cation-exchange chromatographic media Streamline SP for separation and purification of anti-HBsAg Fab fragment from E. coli., Methods: The adsorption of the target protein for separation and purification by the cation-exchange chromatographic media Streamline SP was tested using test tube method in balanced buffer solution with different pH values and ion concentrations. The adsorption effect was then verified by cation-exchange chromatography using 1-ml Streamline SP prepacked column and 28-ml Streamline SP self-assembly column., Results: According to the experiment results of test tube method, the loading buffer with pH of 4.4 and ionic concentration of 100 to 600 mmol/L could achieve optimal target protein adsorption effect by cation-exchange chromatographic media Streamline SP, as verified by cation-exchange chromatography with 1-ml SP prepacked column and 28-ml Streamline SP self-assembly column., Conclusion: The optimal condition of cation-exchange chromatography selected by test tube method can be applied for separation and purification of anti-HBsAg Fab fragment from E. coli.

In order to produce recombinant human anti-HBsAg Fab antibody in Pichia pastoris, the recombinant yeast was fermented using fed-batch system in a 30 L bioreactor. The fermentation temperature was 30 degrees C, the pH was 5.0 approximately 5.3, and the DO was 20% approximately 30%. The recombinant Fab antibody was purified from crude culture supernatant by ion exchange and analyzed by SDS-PAGE and western blot and ELISA. When the absorbance (OD600) of broth reach 300 at the end of fed-batch phase, the induced phase was initiated. The results showed that recombinant human anti-HBsAg Fab antibody was high-level expressed in recombinant Pichia pastoris using a fed-batch fermentation system. Both chains of the Fab were successfully expressed upon methanol induction. After 192 h of induction, the expression level of recombinant Fab (soluble) reached 412 mg/L. The recombinant Fab antibody was purified effectively by ion-exchange chromatography from the fermentation supernatant to a purity of 95%. And the affinity activities of the purified recombinant Fab antibdy and fermentation supernatant were detected, and both of them showed high affinity activities. The results demonstrated that recombinant human anti-HBsAg Fab antibody could be high level produced by fed-batch fermentations in Pichia pastoris. Which can be efficiently used in industrial production.

Aim: To establish a steady purification method for producing recombinant humanized anti-HBsAg Fab from yeast fermentation supernatant., Methods: Humanized anti-HBs scFv was used to immunize BALB/c mice to obtain anti-scFv monoclonal antibody (mAb) which was used to purify recombinant humanized anti-HBsAg Fab from yeast fermentation supernatant by affinity chromatography., Results: The purity of purified recombinant anti-HBsAg Fab was above 95% and its recovery rate was about 75%-85%., Conclusion: An efficient affinity chromatography suitable for purification of recombinant humanized anti-HBsAg Fab in large-scale was established.

We estimated the prevalence of markers of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection, and injecting risk behaviour, among community-recruited injecting drug users (IDUs) in North West Wales in 2001 and 2002. Sample collection was undertaken by trained current and former IDUs. Oral fluid samples (n = 153) were tested as part of the Unlinked Anonymous Prevalence Monitoring Programme ongoing survey of IDUs. Approximately 12% of the sample reported that they were currently in a drug treatment programme. Of the 153 samples screened 27% (95% CI 20%-34%, 41/153) were anti-HBc positive, and 23% (95% CI 16%-30%, 35/153) were anti-HCV positive. Sixteen per cent (95% CI 10%-22%, 25/ 153) of the samples were positive for both anti-HBc and anti-HCV. Of the subjects 15% (95% CI 9%-20%) knew they had been vaccinated against hepatitis B. Direct sharing of needles and syringes in the 28 days prior to interview was reported by 44% (95% CI 35%-54%), and sharing of any equipment including that used for drug preparation prior to injection was reported by 66% (95% CI 57%-76%). In North West Wales, syringe sharing is a common practice, and a high proportion of IDUs have been exposed to bloodborne viruses. Hepatitis B vaccination coverage within this population appears to be low and needs to be increased. Further efforts are needed to improve the availability of clean injecting equipment.

The recombinant human monoclonal antibody (MAb) against hepatitis B virus (HBV) surface antigen (HBsAg) was expressed in tobacco suspension cultures. The parental CL4MAb was produced by the Epstein-Barr virus (EBV) transformed human cell line TAPC301-CL4. The CL4MAb cDNA was introduced into tobacco suspension cells by Agrobacterium mediated transformation. The monoclonal antibodies (MAbs), B294 and B303, which were derived from CL4 and subsequently produced in plant cells were selected for study. After purification on Protein A columns, B294 and B303 MAbs had anti-HBs relative affinity constants similar to the parental CL4MAb. B303 MAb interacted with cell surface HBsAgs and showed complement-dependent cytotoxicity in a manner that was similar to anti-HBs human immunoglobulins (HBIg) that are used clinically. The results of this study point to the feasibility of producing MAbs to HBsAg in plants as an alternative to HBIg., (Copyright 2004 Wiley-Liss, Inc.)

Background and Objectives: Nanofiltration is one of the most effective virus reduction methods in the manufacturing process of plasma products. However, it is difficult to remove small viruses from high molecular weight protein preparations like immunoglobulin G or factor VIII complex by nanofiltration, because the size of the protein is similar to that of viruses. In order to separate the viruses from these proteins by nanofiltration, it is necessary to change the size of either one. In this study, we report that such non-enveloped viruses as human parvovirus B19 (B19), human encephalomyocarditis virus (EMC) or porcine parvovirus (PPV) aggregate in the presence of certain kinds of amino acids and could be easily removed by nanofiltration., Materials and Methods: 0.3 M Glycine (or other amino acid) solution spiked with viruses was subjected to dead-end single filtration with a 35-nm pore-size filter. Virus removal by nanofiltration was either evaluated by PCR or by infectivity assay., Results: B19 in a 0.3 M glycine solution was reduced to 1:10(7.5) (7.5-log) by nanofiltration with a 35-nm pore-sized filter, whereas in PBS it was not reduced. Similarly, B19 was also reduced when suspended in other amino acids solutions. This effect was also confirmed with the other small non-enveloped viruses EMC or PPV. When 5% globulin or 5% albumin was added to a 0.3 M glycine solution, the removal rate was decreased., Conclusions: These data suggest that viruses in the presence of certain kinds of amino acids could be aggregated and effectively removed by a filter that has a pore size larger than the size of the viruses.

Anti-HBs Fab fragment has considerable potential for use in the prevention and treatment of liver diseases by HBV. Here we established a high-level expression system to directly produce anti-HBs Fab fragment in Pichia pastoris. This was achieved by co-integration of the genes encoding the heavy and light chains both under the genome of the yeast cells. The Fab fragment was efficiently secreted into medium at a concentration of 50 mg/liter. The authenticity of the Fab fragment was confirmed by immunoblot analysis, which yielded one band of approximately 50 kDa under nonreducing conditions and two bands of approximately 28 kDa under reducing conditions. The anti-HBs Fab fragment was prepared with a purity of 95% by affinity chromatography. The affinity activity of the recombinant Fab was detected by ELISA, which indicated that 1 mg of recombinant Fab was equivalent to 40 IU HBIG (20 IU/mg). The results demonstrated that the recombinant Fab fragment could sufficiently neutralize the HBsAg.

Transgenic plants expressing recombinant immunoglobulins have arisen as an alternative technology for the large-scale production of antibodies useful in therapeutics and in industrial processes. In the present paper we report the expression in transgenic tobacco ( Nicotiana tabacum ) of an anti-HBsAg [anti-(hepatitis B virus surface antigen)] mouse IgG1 mAb (monoclonal antibody), currently used for the industrial purification of the recombinant vaccine antigen. Using the sweet potato sporamin signal peptide, a KDEL (Lys-Asp-Glu-Leu) ER (endoplasmic reticulum) anchorage domain, and a heavy- and light-chain gene tandem construction, we generated F1 plants in which the expression of the antibody accounted for 0.5% of the total soluble proteins. The 'plantibody' (functional IgG antibody produced in plants) was easily purified by Protein A-Sepharose chromatography with a yield of approximately 35 microg/g of fresh leaf material, and its glycosylation indicated that, irrespective of the KDEL signal, the molecule is modified in both the ER and Golgi. Finally, a successful comparison of the plantibody with the ascites-derived mAb in the immunoaffinity purification of the vaccine recombinant HBsAg was performed. Taken as a whole, our results show that the large-scale production of this antibody of industrial relevance in transgenic tobacco is feasible.

We screened 651 human immunodeficiency virus (HIV)-1-infected subjects for hepatitis B surface antigen (HBsAg) and antibody to hepatitis B surface antigen (anti-HBs). Of a total of 387 subjects who tested negative for both HBsAg and anti-HBs, 142 underwent further testing for isolated presence of antibody to hepatitis B core antigen (anti-HBc). Of these 142 subjects, 60 (42%) tested positive for anti-HBc (isolated anti-HBc). Individuals coinfected with HIV-1 and hepatitis C virus (HCV) were more likely to have isolated anti-HBc than were subjects with HIV-1 alone (80% vs. 16%, respectively). Our findings suggest that individuals with HIV-1/HCV coinfection for whom there is no serological evidence for hepatitis B virus when screened with HBsAg and anti-HBs will be positive for anti-HBc in >75% of cases. A screening strategy that tests only for HBsAg and anti-HBs in HIV-1-infected patients will miss a large number of individuals with isolated anti-HBc.

The presence of serological markers for hepatitis B virus (HBsAg, anti-HBc IgM and Anti-HBc total) was investigated in the serum of 1,396 individuals who had clinical suspect of hepatitis. It was observed that 50.7% of the individuals were positive and, from the total of the studied individuals, 14.5% were positive for HBsAg. From these, 8.5% were also positive for anti-HBc IgM. The analysis in relation to gender showed a higher seroprevalence index among male individuals (p < 0.0001). It was observed the occurrence of subtypes adw2 (62.7%), ayw3 (23.5%), ayw2 (9.8%) and adw4 (3.9%). The viral DNA was detected in 61 (33.9%) HBsAg positive samples and in one sample positive only for anti-HBc total. These results indicate an important incidence of the HBV infection in this population, and reinforce previous studies regarding this virus in the central west region of Brazil.

Using of two-step integrating technology, transducted the H and L chain gene of humanized Fab fragment of anti-HB-sAg antibody into the genome of methylotropic yeast P. pastoris. Constructed a engineering yeast to produce humanized Fab fragment of the anti-HBsAg antibody. The Fab fragment was efficiently secreted into the medium at a concentration of 50-80 mg/L. The Fab fragment was purified from culturing supernatant of the recombinant yeas by affinity chromatography. The ELISA analysis showed the high affinity of the expressed humanized Fab fragment to the HBsAg.

The infectivity of 310 hepatitis B carriers (among antenatal and genitourinary medicine clinic attendees, blood donors, and patients of a liver disease unit) was assessed using three different assays: chemiluminescent molecular hybridisation assay (Murex Digene), column-based solution hybridisation assay (Abbott Genostics) and in-house polymerase chain reaction (PCR). PCR was found to be at least 100 times more sensitive (1 x 10(4) copies/mL) than Murex Digene (3.2 x 10(6) copies/mL) and Abbott Genostics (3.7 x 10(7) copies/mL). Comparison of the hepatitis B e antigen (HBeAg)/anti-HBe status and hepatitis B virus (HBV) DNA level confirmed an association between these two variables. The overall detection rate of HBV DNA by Murex Digene was 28% (87/310): 89% (73/82) in the HBeAg positive group, 10% (4/40) in the HBeAg/anti HBe negative group, and 5% (10/188) in the anti-HBe positive group. The detection rate by PCR increased to 53% (163/310): 98% (80/82) in the HBeAg positive group, 38% (15/40) in the HBeAg/anti-HBe negative group, and 36% (67/188) in the anti-HBe positive group. HBV DNA detection rates by all three assays in 97 liver disease unit patients were higher, particularly in anti-HBe positive patients, than in the cohort overall, probably reflecting a higher rate of active liver disease in these patients. HBV DNA was detected at the lowest rate in the antenatal clinic group. We suggest that HBeAg negative patients who are positive by PCR but negative by either Murex Digene or Abbott Genostics are still infectious. A cut-off serum HBV DNA level of 10(4) copies/mL is proposed, below which transmission is unlikely to occur, but further studies using quantitative PCR are needed to refine the cut-off level.

Objective: To verify the prevalence of the anti-HBc, anti-HBs and HBsAg markers of hepatitis B virus, and to identify the risk factors determining occupational infection with this virus among hospital personnel., Methods: Samples of serum from 210 persons both male and female who work in different occupations at a hospital university, were analysed. The technique employed was the immunoenzymatic assay using commercial kits., Results: As a control group, samples of serum from 45 volunteer blood donors were utilized. It was verified that 20.5% of the hospital personnel presented a positive reaction to at least one of the markers songht, as against 6.6% of the control group. The prevalence of each marker separately was: anti-HBc 8.1%, anti-HBs 5.2%, and HBsAg 2.9% in the hospital personnel; and 4.4%, 2.2% and 0.0% in the control group. The simultaneous presence of the anti-HBc and anti-HBs markers was detected in 4.3% of the workers. In the control group, the presence of the anti-HBc and anti-HBs markers was detected, isolately, with respective prevalences of 4.4% and 2.2%. Those who presented the highest rates of positivite reaction were: laboratory technicians 24.0%, nurses 23.6%, physicians 20.8%, and cleaning personnel 18.2%., Conclusions: The findings suggest that direct contact with patients and handling of blood and other body fluids are risk factors related to occupational infection with HBV. Therefore, it is recommended that hospital personnel be vaccinated against hepatitis B.

Blood transfusion in the Netherlands depends wholly on voluntary donations of blood. Very soon the measures to ensure the safety, the quality and clinical appropriateness of the application of blood products will be re-evaluated. Currently it is not so much viral but bacterial contamination that causes worry, in addition to immunological transfusion reactions. Safety not only depends on freedom from viral or bacterial contamination, but also on the appropriate clinical use of blood products. Physicians will need to document the individual indications for the use of blood products and the patient's reactions at follow-up. European legal harmonisation emphasises look-back procedures and haemovigilance as a means of ensuring sound clinical practice regarding blood transfusion.

Hemodialized pediatric patients are a risk population for the hepatitis B and C virus infection. The aim of this paper was to study the serum prevalence of HBV and HCV infection in hemodialized children. We study 61 pediatric patients at hemodialisis, 12 on renal transplant, range between 2 and 20 years old (mean: 12.9 years), 23 male and 38 female. The specific anti-HCV IgC were measured by enzyme immunoassay (ELISA Abbott) and confirmed by LIA-TEK (Organon). The anti-HBV were measured by ELISA Abbott and transaminases by cinetic method (ASAT: 29 UI/L and ALT: 33 UI/L). The 19.7% of studied children were HCV (+) and 29.5% were HBV (+), 38.9% of them were HbsAg (+) and 50% anti-HBs (+). The HCV and HBV infection was more elevated in relation to the transfusion number and the hemodilisis time. The elevation of ALT/ASAT activity isn't a right infection index for HCV and HBV in this children.

In this paper we describe the antigen recognition characteristics, variable region base and amino acid sequence, and performance as immunoaffinity chromatography ligands of two MAb specific to the alpha determinant of the HBsAg, derived from the same fusion. We show that the epitope recognized by CB-Hep.0 (IgM) is probably associated to an intrachain disulfide bond in the antigen. On the other hand, CB-Hep.1 (IgG2b) recognizes a heat-resistant non-conformation dependent antigenic determinant on HBsAg. PCR-cloning and sequencing of the variable regions of these two MAb indicated that both heavy chain variable regions were originated from the usage of the same germinal V and J genes. However, the outstanding differences in the size of the VH CDR3, and the absolute difference in the light chain sequences, suggest that the hybridomas were originated from different precursor B lymphocytes. With respect to their use as immunoaffinity chromatography ligands for the purification of a recombinant HBsAg, we found that the IgM immunogel exhibited increased performance with respect to amount of eluted antigen, and final recovery. This difference in overall performance could be attributed to a series of factors: the higher valence number of IgM, a dissimilar distribution of IgM and IgG in the activated gel particles, and differences in antigen recognition between both MAb. Our results suggest that IgM antibodies may be useful in immunopurification, particularly if the antigen is structurally complex and has a high density of repeating epitopes.

For industrial production of human monoclonal antibodies (hmAb) against hepatitis B virus surface antigen (HBsAg), we scaled-up a short-term perfusion culture in serum-free medium, which was chosen as the most suitable culture method, to a 50-1 fermentor equipped with a rotating shear filter. Using hydrophobic chromatography as the initial step of hmAb purification, the mAb HBW4, HBW6 and W471 were isolated in good quality from the respective culture broths in yields of approximately 75%. Each of the three purified hmAb alone, and a cocktail of the three, protected chimpanzees against HB virus, when injected intravenously 3 h after viral challenge, as long as the serum antibody levels were significant. A pharmacokinetic study using cynomolgus monkeys demonstrated that the hmAb have a long plasma half-life and bioavailability of approximately 76% upon intramuscular injection in primates. Thus, anti-HBsAg hmAb produced by an industrial process are expected to be successfully used in clinical fields.

Until today only five pathogens of viral hepatitis have been discovered: hepatitis virus A (HAV), B (HBV), C (HCV), D (HDV) and E (HEV). An update on the state of the art of actually established diagnostic procedures is briefly summarized. Some recent topics and problems are covered more extensively; in the case of HBV the value of HBsAG concentration as a prognostic marker, the finding 'anti-HBc only' the role of HBV mutants, the determination of circulating HBV genomic material, and modalities for HBV vaccination control; in the case of HCV the indications for study of its viral genomic material and the role of HCV subspecies are explained. Finally, first results on HEV prevalence in Switzerland are introduced.

A second generation assay for antibody to hepatitis C virus (anti-HCV) was used in order to establish the exact prevalence of HCV infection in haemodialysis patients. HCV RNA was sought by the polymerase chain reaction in order to determine whether haemodialysis patients with anti-HCV had been infected with HCV in the past or were presently infected. Of 357 patients, 198 (55.5%) were positive for anti-HCV, compared to 113 (31.7%) positive for original antibody to c100-3 protein (P < 0.001). HCV RNA was detected in 171 (86.4%) of the 198 patients with anti-HCV. Liver dysfunction was found in 63 (17.6%) of all haemodialysis patients. Of these, 55 (87.3%) had HCV infection, one (1.6%) hepatitis B virus infection while, in the remaining seven, the origin was unknown. Thus, in almost all anti-HCV-positive patients, HCV viraemia was present. We conclude that HCV is an important cause of liver disease in haemodialysis patients.