Detection of a hepatitis B surface antigen variant emerging in a patient with chronic lymphocytic leukaemia treated with fludarabine

Fludarabine is used widely for the treatment of chronic lymphocytic leukaemia, but not as yet implicated in the emergence of hepatitis B surface antigen (HBsAg) variants following hepatitis B virus (HBV) reactivation. Such a variant was detected in a 78-year-old female who was HBsAg(-)/anti-HBc(+)/anti-HBs(+)/anti-HBe(+), and with normal ALT levels, who developed HBV reactivation after fludarabine treatment. She had high HBV-DNA levels, and became positive for HBeAg, in the absence of detectable HBsAg. HBV-DNA was extracted from serum and the HBsAg encoding region of the genome was amplified by PCR, followed by cloning and sequencing. The HBV strain appeared to be subtype adw, but had higher nucleotide homology with ayw than adw isolates, supported further by phylogenetic tree analysis. Amino-acid sequence comparisons over the alpha determinant region revealed the following substitutions: C124N, G130R, and N146S. There were also unique substitutions outside the alpha determinant. All these mutations appeared to have a profound effect on the antigenicity of this region, which resulted in failure to detect HBsAg by commercially available diagnostic assays. It is concluded that a surface variant emerged in an HBsAg(-)/anti-HBs(+) patient with chronic lymphocytic leukaemia following fludarabine treatment, with an unprecedented number of amino-acid substitutions in the alpha determinant region of HBsAg, including a subtype switch.     

Detection of a premature stop codon in the surface gene of hepatitis B virus from an HBsAg and antiHBc negative blood donor.

BACKGROUND: In blood donors, HBV infection is detected by the presence of serum hepatitis B surface antigen (HBsAg). However, some mutations in the surface gene region may result in altered or truncated HBsAg that can escape from immunoassay-based diagnosis. Such diagnostic escape mutants pose a potential risk for blood transfusion services. RESULTS: In the present study, we report a blood donor seronegative for HBsAg and antiHBc, but positive for antiHBs who was HBV DNA positive by PCR. Sequencing of the HBsAg gene revealed presence of a point mutation (T-A) at 207th nucleotide of the HBsAg ORF, which resulted in a premature stop codon at position 69. This results in a truncated HBsAg gene lacking the entire 'a' determinant region. However, follow-up of the donor after 2 years revealed clearance of HBV DNA from the serum. CONCLUSION: The case illustrates an unusual mutation, which causes HBsAg negativity. The finding emphasizes the importance of molecular assays in reducing the possibility of HBV transmission through blood transfusion. However, developing more sensitive serological assays, capable of detecting HBV mutants, is an alternative to expensive and complex amplification-based assays for developing countries.     

Detection of hepatitis B virus DNA in the serum of Canadian hepatitis B surface antigen negative,anti-HBc positive individuals,using the polymerase chain reaction

Continuing hepatitis B virus (HBV) infection is normally associated with the presence of hepatitis B surface antigen (HBsAg) in the serum. In spite of sensitive screening assays for HBsAg, rare cases of post-transfusion HBV infection are still observed. Antibody to hepatitis B core antigen (anti-HBc) often indicates remote HBV infection but DNA hybridisation and more sensitive polymerase chain reaction (PCR) assays have demonstrated that some HBsAg negative individuals, positive for anti-HBc, have continuing HBV replication. To determine the incidence of ongoing HBV infection in a Canadian HBsAg negative, anti-HBc positive population we studied three groups with this combination of HBV markers: Group A, 36 patients referred for investigation of raised serum aminotransferases; Group B, 21 Canadian Red Cross blood donors; Group C, seven vaccinees in an Ottawa Health Care Student hepatitis B vaccination programme. The PCR was carried out using a nested PCR reaction with primers specific for the pre-core region of HBV. Seven of 36 (19%) patients in Group A had detectable HBV DNA whereas none of Group B or C were positive. This data indicates that in some HBsAg negative patients with ongoing hepatic inflammation, continuing HBV replication may persist. This was not observed in any healthy blood donors or health care students investigated. Larger studies are required, but this data would suggest that, in Canada, the addition of anti-HBc testing for all blood donors for detection of low level HBV replication would not be indicated. © 1994 Wiley-Liss, Inc.     

Improved Detection of Hepatitis B Virus Surface Antigen by a New Rapid Automated Assay

The performance of hepatitis B virus (HBV) surface antigen (HBsAg) screening assays is continuously improved in order to reduce the residual risk of transfusion-associated hepatitis B. In a multicenter study, a new automated rapid screening assay, Elecsys HBsAg (Roche Diagnostics), was compared to well-established tests (Auszyme Monoclonal [overnight incubation] version B and IMx HBsAg [Abbott]). Included in the evaluation were 23 seroconversion panels; sera from the acute and chronic phases of infection; dilution series of various HBsAg standards, HBV subtypes, and S gene mutants; and isolated anti-HBV core antigen-positive samples. To challenge the specificity of the new assay, sera from HBsAg-negative blood donors, pregnant women, and dialysis and hospitalized patients and potentially cross-reactive samples were investigated. Elecsys HBsAg showed a higher sensitivity for HBsAg subtypes ad, ay, adw2, adw4, ayw1, ayw2, ayw4, and adr detection in dilution series of different standards or sera than Auszyme Monoclonal version B and/or IMx HBsAg. Acute hepatitis B was detected in 11 to 16 of 23 seroconversion panels between 2 and 16 days earlier with Elecsys HBsAg than with the alternative assays. Elecsys HBsAg and Auszyme Monoclonal version B detected HBsAg surface mutants with equal sensitivity. The sensitivity and specificity of Elecsys HBsAg were 100%. Auszyme Monoclonal version B had a 99.9% specificity, and its sensitivity was 96.6%. IMx HBsAg showed a poorer sensitivity and specificity than the other assays. In conclusion, Elecsys HBsAg permits earlier detection of acute hepatitis B and different HBV subtypes than the alternative assays. By using highly sensitive HBsAg screening assays, low-level HBsAg carriers among isolated anti-HBV core antigen-positive individuals can be detected.     

Validity of the rapid strip assay test for detecting HBsAg in patients admitted to hospital in Uganda

Commercially available rapid strip assays (RSAs) for hepatitis B surface antigen (HBsAg) are used for most routine clinical testing in sub‐Saharan Africa. This study evaluated the validity of RSA and a more sophisticated enzyme immunoassay (EIA) with confirmation by nucleic acid testing (NAT) in hospitalized patients in Uganda. Sera from 380 consecutive patients collected and tested for HBsAg and anti‐HIV in Kampala, Uganda by RSA were sent frozen to Dallas for EIA including HBsAg, total anti‐hepatitis B core, hepatitis B e antigen, and anti‐HIV. NAT was performed on all HBsAg‐positives and on a random sample of 102 patients that were HBsAg‐negative by both assays. Overall, 31 (8%) were HBsAg positive by RSA while 50 (13%) were HBsAg‐positive by EIA; 26 were concordant between the two assays. Of 55 HBsAg‐positive patients, nearly all showed detectable serum hepatitis B virus (HBV) DNA by bDNA (46) or PCR (4) assay. The 26 patients who were HBsAg positive by both EIA and RSA had significantly higher median serum HBV DNA levels than the 24 patients who were HBsAg positive by EIA alone. An additional 12/102 (12%) HBsAg negative patients had very low serum HBV DNA levels by NAT. Several differences in expected results of serologic testing were observed in this large series of African patients. RSA HBsAg testing is less sensitive than EIA; even EIA failed to detect all HBV DNA positive sera. A more complex testing protocol than RSA alone will be needed in Africa to improve patient care. J. Med. Virol. 82:1334–1340, 2010. © 2010 Wiley‐Liss, Inc.     

False-Negative Hepatitis B Virus (HBV) Surface Antigen in a Vaccinated Dialysis Patient with a High Level of HBV DNA in the United States

Screening with hepatitis B surface antigen (HBsAg) is highly recommended for at-risk individuals. Mutations in the HBsAg can result in an inability to detect the virus during routine screening. We describe a hemodialysis patient found to have high levels of hepatitis B virus (HBV) DNA and HBV antibody but negative HBsAg on two routine assays.     

Hepatitis B surface antigen containing immune complexes occur in seronegative hepatocellular carcinoma patients

IgG, IgM and hepatitis B surface antigen (HBsAg) containing immune complexes (IC) were detected by the Clq and conglutinin solid phase assays in both HBsAg+ and HBsAg- groups of patients with primary hepatocellular carcinoma (HCC). No differences were observed between the two patient groups either in the levels of antigen non-specific and HBsAg specific complexes or in the immunoglobulin isotype in the complexes. The results show that HBsAg can occur in an IC form in the sera of patients classified as HBsAg- by sensitive commercial assays and provides evidence of a further association of hepatitis B virus (HBV) and HCC in antigen negative patients. Furthermore, the HBsAg IC in HCC patients differ from those in other HBV infected subjects in that they are preferentially detected by the Clq assay.     

Naturally occurring MHR variants in Turkish patients infected with hepatitis B virus

Major B-cell epitopes are located at the major hydrophilic region (MHR) of hepatitis B virus (HBV) surface antigen (HBsAg). The genotypes, subtypes, and naturally occurring amino acid (aa) substitutions of MHR were analyzed in 81 Turkish adult patients (41 inactive HBsAg carriers and 40 patients with chronic hepatitis B) by direct sequencing of the S gene fragment. All the isolates were genotype D according to the phylogenetic analysis. The most common HBsAg subtype was ayw2, followed by ayw3 while one isolate specified ayw4 by encoding Leu127. MHR variants were detected in 22 of the 81 (27.2%) isolates. The prevalence was significantly higher in the chronic hepatitis B group (42.5%) compared to inactive HBsAg carriers (12.2%). Twenty-two samples had a total of 26 amino acid substitutions involving 14 positions. The majority of the patients had a single variation. Most of the amino acid substitutions were located at the HBs1 region of the MHR, while 9 of the 26 were in the classic "a" determinant (aa 124-147). When samples with "a" variants were evaluated by two different commercial HBsAg tests, only the isolate with Ser143Leu variation had a decreased reactivity in the assay using monoclonal antibodies for capture and detection. In conclusion, the findings of the study was in accordance with previous studies showing HBV genotype and subtype homogeneity (genotype D/ayw) in Turkey. Naturally occurring MHR and "a" determinant variants were common, especially among chronic hepatitis B patients. The influence of detected "a" variants on diagnostic assays was limited.     

Reactivation of hepatitis B virus infection with persistently negative HBsAg on three HBsAg assays in a lymphoma patient undergoing chemotherapy

In patients with occult hepatitis B virus (HBV) infection, acute exacerbation may occur when they become immunocompromised. Usually, these patients develop hepatitis B surface antigen (HBsAg) seroreversion during the flare. Here we report on a patient with occult HBV infection, who developed HBV exacerbation after chemotherapy for diffuse large B-cell lymphoma. The resurgence of HBV DNA preceded the elevation of liver enzymes for 20 weeks. Atypically, despite high viraemia, serological tests showed persistently negative HBsAg using three different sensitive HBsAg assays (i.e., Architect, Murex and AxSYM). On comparing the amino acid sequence of the index patient with the consensus sequence, five mutations were found at pre-S1, five at pre-S2 and twenty-three mutations at the S region. Six amino acid mutations were located in the ‘a’ determinant, including P120T, K122R, M133T, F134L, D144A and G145A. The mutants K122R, F134L and G145A in our patient have not been tested for their sensitivity to Architect and Murex assays by the previous investigators and might represent the escape mutants to these assays.     

Significance of anti-HBc IgM in the differential diagnosis of viral hepatitis

IgM antibody to hepatitis B core antigen (anti-HBc IgM) as determined by IgM capture immunoassay is generally present in high titer during acute hepatitis B infection. A strong positive reaction for anti-HBc IgM during acute hepatitis is indicative of an acute HBV infection even in hepatitis B surface antigen (HBsAg)-negative patients. With the help of anti-HBc IgM otherwise unidentified HBV infection can be diagnosed in HBsAg-negative patients and an optimal combination of diagnostic tests for acute hepatitis B infection would therefore include assays for both HBsAg and anti-HBc IgM. In the HBsAg carrier with or without chronic liver disease the presence and meaning of anti-HBc IgM is still a matter for discussion. Detection of a weak positive result for anti-HBc IgM in HBsAg-positive patients without a recent history of acute hepatitis cannot always be regarded as a definite marker of recent hepatitis B infection. However. quantitation of the anti-HBc IgM results seems to improve the clinical value of the test. Comparison of the available anti-HBc IgM assays is needed and may well establish a reliable cut-off level that would differentiate acute from chronic hepatitis B and ongoing from resolving hepatitis B in HBsAg-positive patients.     

Prevalence of hepatitis B virus MHR mutations and their correlation with genotypes and antiviral therapy in chronically infected patients in Serbia

Understanding the prevalence and diversity of HBsAg variants in a population is fundamental to assay design and planning vaccination programs. It has been shown that mutations within the S gene, caused by selection or natural variation, can lead to false‐negative results in assays for HBsAg, or have clinical implications, such as evading anti‐HBV immunoglobulin therapy or vaccine‐induced immunity. The region of HBsAg where most of these mutations occur is known as the major hydrophilic region (MHR). The aim of this study was to determine the prevalence and mutational patterns of MHR mutations in patients with chronic hepatitis B, and their correlation with patient characteristics, viral factors and antiviral therapy. The study comprised 164 plasma samples from patients with chronic hepatitis B, of which, 34.8% were on long‐term lamivudine monotherapy. Direct sequencing of part of the S/pol gene was used for identification of HBsAg mutations, HBV genotypes, subgenotypes and HBsAg subtypes. The overall frequency of MHR mutations was 22.6%, but it varied significantly between untreated and treated patients (16.8% vs. 33.3%). The most frequent substitution was at position 120 (9.1%) whereas the most common vaccine‐escape position, 145, was affected in 1.8% of isolates. The presence of MHR mutations was correlated with genotype D, subgenotype D3, and ayw2/ayw3 HBsAg subtypes and to older age (>40 years). It is concluded that natural viral variability present in a geographical region, duration of infection, and antiviral therapy are among the major factors associated with the occurrence of MHR mutations. J. Med. Virol. 82: 1160–1167, 2010. © 2010 Wiley‐Liss, Inc.     

Cell-mediated immunity to hepatitis B surface antigen in man.

An aqueous preparation of hepatitis B virus (HBV) vaccine was used as an intradermal skin test antigen to assess delayed hypersensitivity to hepatitis B surface antigen (HBsAg). Thirty-five persons were tested including 10 individuals seronegative for all HBV markers, 10 positive for HBsAg (chronic carriers) and 15 positive for antibody to HBsAg (anti-HBs), five of whom had received the HBV vaccine. All patients were also studied for lymphocyte blastogenic responses to phytohaemagglutinin, concanavalin A, pokeweed mitogen and purified HBsAg. Only one individual had a positive delayed skin test reaction to HBsAg. This person had received the HBV vaccine and had high titres of anti-HBs in serum. However, neither this individual nor any other subject exhibited a positive lymphocyte blastogenic response to HBsAg in vitro. Thus, delayed hypersensitivity skin test reactivity to HBsAg was not detected after natural infection with HBV and was rarely present in hyperimmunized individuals. In vitro assays of immune responsiveness failed to demonstrate cellular immunity to HBsAg even in hyperimmunized persons. These studies provide no evidence that cell-mediated immunity to HBsAg plays a role in the immunopathogenesis of acute or chronic type B hepatitis.     

Sensitivities of four new commercial hepatitis B virus surface antigen (HBsAg) assays in detection of HBsAg mutant forms

Mutations in hepatitis B virus surface antigen (HBsAg) involving amino acid substitution within the immunodominant "a" determinant may affect the performance of commercial HBsAg assays. The performances of four HBsAg assays that recently received Conformité Européene marking, Advia Centaur HBsAg (Bayer), Monolisa HBsAg Ultra (Bio-Rad), Liaison HBsAg (Dia Sorin), and Vidas HBsAg Ultra (bioMérieux), were compared with that of the routinely used HBsAg assay AxSYM HBsAg V2 (Abbott). Assays were evaluated for (i) analytical sensitivity performance with a national reference HBsAg panel (including 10 samples with calibrated HBsAg concentrations from 0.04 to 2.24 ng/ml) and (ii) the detection of HBsAg mutants by studying a panel of 35 HBsAg mutants (23 collected from patients and 12 recombinant mutants). The limits of detection of these assays were <0.15 ng/ml (from 0.089 to 0.121 ng/ml). The sensitivity performances for mutant virus detection varied, ranging from 37.1% to 91.4%. The lack of detection of these mutants by commercial assays was probably due to the epitope recognition of the anti-HBs assay reagents in the capture phase and in the conjugates. The prevalence and clinical impact of HBsAg mutants are under investigation. However, the manufacturers must be vigilant in the design of the assays in order to reduce the risk of missing a broad range of described S gene mutants.     

Correlation of HBV DNA and monoclonal reactivity to HBsAg in serum of patients with HBV infection

Hepatitis B virus (HBV) DNA hybridization assay, a monoclonal radioimmunoassay (M-RIA) for hepatitis B surface antigen (HBsAg) and conventional polyclonal immunoassays for HBV associated antigens were used to study sera from patients on dialysis and with acute hepatitis B. HBV DNA was detectable in hepatitis B e antigen (HBeAg) negative patients with acute hepatitis but not in HBsAg+ HBeAg- dialysis patients. In acute hepatitis, HBsAg immunoreactivity by M-RIA could still be detected even though a commercial immunoassay for HBsAg, the AUSRIA II, and the HBV DNA assay were no longer positive. Unlike in acute HBV infection, serum HBV DNA was detectable in dialysis patients who were AUSTRIA II negative but M-RIA positive. Serial determination of HBsAg by M-RIA and HBV DNA revealed episodes of HBV DNA positivity months after both the HBsAg was no longer positive by polyclonal immunoassay. Thus, the M-RIA for HBsAg and the molecular hybridization technique for HBV DNA are sensitive and specific assays for the identification of potentially infectious individuals who would not have been characterized as such based on the results of conventional polyclonal immunoassays.     

Differential diagnosis of acute viral hepatitis using rapid, fully automated immunoassays

We report the development of three rapid, fully automated immunoassays allowing the differential diagnosis of acute viral hepatitis. These assays detect HBsAg, IgM antibody to hepatitis B core antigen (IgM anti-HBc) and IgM antibody to hepatitis A virus (IgM anti-HAV) using the IMx instrument system. All IMx assays were run in less than 45 minutes and all steps were fully automated including specimen dilution steps. Specimens from blood donors, diagnostic and hospital patients, and individuals with a variety of infectious and immune diseases were tested for IgM anti-HAV (n = 1473) or for IgM anti-HBc (n = 1606) or for HBsAg (n = 9700) by the IMx and commercially available EIA and RIA. Each IMx assay showed 99.8% agreement with current EIA. Reproducibility in all hepatitis IMx assays was significantly better than that observed with manual or semiautomated assays; within-run and between-run % CV ranged from 2.2 to 4.8 and 3.5 to 10.3 respectively. In 29 acute hepatitis B patients studied, HBsAg and IgM anti-HBc were detected in the first available patient bleed collected from 0 to 4 week from the onset of symptoms. IgM anti-HBc persisted at reactive levels in the IMx assay for 1 to 24 weeks (mean 12.1 +/- 5.3 weeks) after the patient presented with symptoms. In individuals exposed to hepatitis A, IgM anti-HAV was detectable by IMx by 40 days post exposure (average 33.5 days) and IgM had declined to unreactive levels in IMx for all patients by from 3 to 6 months post exposure. These data demonstrate the use of these rapid IMx assays for differentiation of acute hepatitis A and B.     

乙肝表面抗原诱生γ-干扰素的研究

对 8名抗 HBs阳性健康成人 ,12名慢性乙肝患者外周血单个核细胞 (PBMC )分别用酵母菌表达的重组HBsAg ,s抗原第 12 0～ 147位氨基酸合成肽及重组HBsAg HBIG复合物刺激 ,测定特异性诱生γ 干扰素水平。结果 2 0名中有 14名产生了比对照更高的γ 干扰素。用重组HBsAg与合成肽为抗原诱生的γ 干扰素阳性数基本相符 ,但用HBsAg HBIG复合物诱生 ,仅有 7名产生升高的γ 干扰素。结果提示今后可用酵母菌表达的重组HBsAg特异诱生PBMC产生γ 干扰素 ,作为一种筛选病人与预测治疗性疫苗疗效的指标     

Sites that bind polymerized albumin on hepatitis B surface antigen particles: detection by radioimmunoassay.

Antibodies to polymerized human albumin (poly-HSA) could not be detected by using sensitive methods (enzyme-linked immunosorbent assay and radioimmunoprecipitation) in sera from chronic carriers of hepatitis B surface antigen (HBsAg) or in serial bleedings from one chimpanzee infected with type A hepatitis virus and one infected with non-A, non-B hepatitis virus. By a solid-phase radioimmunoassay, receptor sites for poly-HSA could be detected on HBsAg particles from sera containing either hepatitis B "e" antigen (HBeAg) or anti-HBe. Blocking experiments showed that monomeric HSA did not bind to this receptor. In general, the HBsAg particles from sera with HBeAg had more poly-HSA receptor sites or relatively more particles carrying this receptor compared with HBsAg from sera with anti-HBe. Microtiter plates coated with poly-HSA bound HBsAg from sera containing HBeAg with greater efficiency than did anti-HBs coupled to a solid phase (Ausria II beads), whereas with sera positive for anti-HBe, the two assays were equally sensitive. Decreased ability of HBsAg to bind to poly-HSA was seen in some sera which had been stored for a few years at 4 degrees C, whereas the binding to anti-HBs was unaffected. It is possible that polymers of albumin on the surface of hepatocytes could function as receptors for hepatitis B virus.     

The amino Acid residues at positions 120 to 123 are crucial for the antigenicity of hepatitis B surface antigen

The major hydrophilic region (MHR) of hepatitis B surface antigen (HBsAg) harbors conformational B-cell epitopes and is the major target of neutralizing antibodies to HBsAg (anti-HBs). Mutant HBsAg (mtHBsAg) with amino acid substitutions such as G145R is known to affect the binding of specific anti-HB antibodies and their detection by conventional diagnostic assays. In the present study, we focused on the role of the amino acid positions 120 to 123, which are around MHR 2 according to the spectrum of recently identified, naturally occurring mtHBsAg. Strikingly, the amino acid substitution K122I abolished the reactivity of HBsAg in all immunoassays tested so far. Also, mtHBsAg G145R could be clearly detected with four different enzyme-linked immunosorbent assays that were based on monoclonal anti-HB antibodies (MAbs) with high affinity. Positive immunofluorescence staining of mtHBsAg K122I was achieved only by polyclonal anti-HBs, while all MAbs tested failed. mtHBsAg T123N showed a low reactivity in immunoassays and appeared to be secretion defective. The amino acid substitution P120T reduced the binding of anti-HBs but did not completely prevent the detection of mtHBsAg by anti-HB MAbs. The testing of naturally occurring mtHBsAg confirmed that the presence of amino acid substitutions within the region of 120 to 123 is strongly associated with impaired detection in immunoassays. In conclusion, MHR 2 is essential for HBsAg antigenicity, a fact that has not been recognized before.     

Comparison between Elecsys HBsAg II and Architect HBsAg QT Assays for Quantification of Hepatitis B Surface Antigen among Patients Coinfected with HIV and Hepatitis B Virus

Hepatitis B surface antigen (HBsAg) quantification has been steadily gaining interest as a clinical marker of therapeutic efficacy, for which two commercial assays are currently available: Architect HBsAg QT (Architect) and Elecsys HBsAg II (Elecsys). HBsAg quantification was evaluated using both assays in 126 human immunodeficiency virus (HIV) and hepatitis B virus (HBV)-coinfected patients initiating treatment with tenofovir dipivoxil fumarate. Linear regression and correlation were used to establish the relationship between the two methods. Bland-Altman analysis was performed to determine mean between-assay difference and limits of agreement (LOA) (±2 standard deviations [SD]) both overall and stratified on HBV (hepatitis B envelope antigen [HBeAg] status, replication, genotype, HBV mutants) or HIV (CD4⁺ cell count) cofactors. There was a significant correlation between Elecsys and Architect assays (correlation coefficient, r = 0.959; P < 0.001). HBsAg quantification using the Elecsys assay was on average 0.200 log₁₀ IU/ml (LOA, −0.500, 0.800) higher than that using Architect, which was consistent across levels of CD4⁺ cell count, presence of precore and YMDD mutations, and HBeAg status. A slightly larger mean between-assay difference was observed with genotypes A and G (0.196 and 0.201, respectively) versus HBV genotypes D and E (0.036 and 0.030, respectively). Mutations on the S region at position s120/s145 were the only determinant in which the mean between-assay difference in HBsAg quantification was lower than the null value (−0.078). In conclusion, the Elecsys assay, with automatic on-board dilution, is capable of quantifying serum HBsAg levels in HIV-HBV-coinfected patients, with very high correlation with the Architect assay.     

HBsAg diagnostic kits in the detection of hepatitis B virus mutation within "a" determinant

The performance of currently available hepatitis B surface antigen (HBsAg) commercial kits was analyzed by using a panel of 212 well-characterized plasma donors all over the country and a panel of nine recombinant HBsAg mutants containing single point or combinations of mutations between amino acid residues 124 and 147 of the "a" determinant. HBsAg commercial kits in this study were machine-based immunoassays with a one-step sandwich ELISA method using either an automatic closed system or manual system. The sensitivity of all machine-based assays evaluated with 105 HBsAg plasma panels was 100% (95% CL = 95.6-99.9%), whereas the specificity with 107 HBsAg negative plasma ranged from 99.07% to 100% (95% CL = 94.2-99.9%). The relative performance of these kits to detect the hepatitis B virus (HBV) mutant panel members of the "a" determinant was found to differ. Interestingly, any commercial kits with monoclonal antibody capture and polyclonal antibody detection (mono/poly), but not mono/mono Ab capture and detection, could pick up the common HBsAg Gly145Arg mutant either solely or in combination with other mutations within the "a" determinant. New versions of HBsAg test kits should recognize multiple HBsAg epitopes in order to detect mutant HBsAg, together with providing good analytical sensitivity and specificity, because of the importance of these assays in HBV diagnosis and in protecting the safety of the blood supply.