Hepatitis B surface antigen (HBsAg) derived from yeast cells (Hansenula polymorpha) used to establish an influence of antigenic subtype (adw2, adr, ayw3) in measuring the immune response after vaccination

Heijtink, R. A.; Bergen, P. van; Melber, Karl; Janowicz, Zbigniew A.; Osterhaus, Albert D. M. E.

doi:10.1016/s0264-410x(02)00145-7

Cited by 31 publications

(21 citation statements)

References 11 publications

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“…Even in the case of hepatitis B virus, where the a determinant region of hepatitis B virus surface antigen has been deemed immunodominant because of the high rate of vaccine escape mutants observed with altered sequence in this 20-amino-acid-residue stretch (55), antibodies that target multiple regions of the capsid have been postulated. Indeed, when sera from health care workers immunized with various commercially available hepatitis B virus vaccines were assayed against three different HBsAg variants, induced antibodies were able to bind heterologous HBsAg proteins, albeit at reduced levels, as seen in this study, suggesting that antibodies which recognize other regions of hepatitis B virus were present (22).…”

Section: Vol 79 2005 Epitopes On Human Papillomavirus 6 L1 Capsomersupporting

confidence: 56%

Humoral Immune Response Recognizes a Complex Set of Epitopes on Human Papillomavirus Type 6 L1 Capsomers

Orozco

Carter

Koutsky

et al. 2005

J Virol

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Although epitope mapping has identified residues on the human papillomavirus (HPV) major capsid protein (L1) that are important for binding mouse monoclonal antibodies, epitopes recognized by human antibodies are not known. To map epitopes on HPV type 6 (HPV6) L1, surface-exposed loops were mutated to the corresponding sequence of HPV11 L1. HPV6 L1 capsomers had one to six regions mutated, including the BC, DE, EF, FG, and HI loops and the 139 C-terminal residues. After verifying proper conformation, hybrid capsomers were used in enzyme-linked immunosorbent assays with 36 HPV6-seropositive sera from women enrolled in a study of incident HPV infection. Twelve sera were HPV6 specific, while the remainder reacted with both HPV6 and HPV11 L1. By preadsorption studies, 6/11 of these sera were shown to be cross-reactive. Among the HPV6-specific sera there was no immunodominant epitope recognized by all sera. Six of the 12 sera recognized epitopes that contained residues from combinations of the BC, DE, and FG loops, one serum recognized an epitope that consisted partially of the C-terminal arm, and three sera recognized complex epitopes to which reactivity was eliminated by switching all five loops. Reactivity in two sera was not eliminated even with all six regions swapped. The patterns of epitope recognition did not change over time in women whose sera were examined 9 years after their first-seropositive visit.

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Section: Vol 79 2005 Epitopes On Human Papillomavirus 6 L1 Capsomersupporting

confidence: 56%

Humoral Immune Response Recognizes a Complex Set of Epitopes on Human Papillomavirus Type 6 L1 Capsomers

Orozco

Carter

Koutsky

et al. 2005

J Virol

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“…Purified protein constituted only one homogeneous band of~24 kDa (Fig. 4a), which was similar to the values obtained from native HBsAg, HBsAg expressed in mammalian cells, and S. cerevisiae [21,22], and correspondent to the band identified in crude lysis. The yield of recovery of purified HBsAg was 72.5%.…”

Section: Purification Of Recombinant Hbsagsupporting

confidence: 79%

“…Recombinant HBsAg S gene has been expressed previously in the systems such as E. coli; yeast systems including S. cerevisiae, Hansenula polymorpha, P. pastoris, Yarrowia lipolytica; and mammalian cells [6,[8][9][10][21][22][23]. However, the certain limitations restrict the expression of heterologous protein in mammalian cells and in S. cerevisiae.…”

Section: Discussionmentioning

confidence: 99%

Expression, Purification, and Characterization of Hepatitis B Virus Surface Antigens (HBsAg) in Yeast Pichia Pastoris

Liu

Lin

Sun

et al. 2009

Appl Biochem Biotechnol

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Prevention of the prevalence of HB depends upon the development of efficient diagnostic reagent and preventive vaccine. Pichia pastoris offers many advantages over the other expression systems in the production of recombinant HBsAg. In this study, we reported that the recombinant P. pastoris strains were cultured in shake flasks and then scaled up in a 5.0-l bioreactor: approximately 27 mg/l of the protein and the maximal cell OD at 600 nm of 310 were achieved in the bioreactor. The recombinant HBsAg was purified by three steps of purification procedures. SDS-PAGE showed that the purified recombinant HBsAg constituted only one homogeneous band of approximately 24 kDa. CsCl density gradient ultracentrifugation assay indicated that the density of the HBsAg was 1.2 mg/ml, which was in agreement with the natural HBsAg, the HBsAg expressed in Saccharomyces cerevisiae and in mammalian cells. Electron microscope observation revealed that the purified recombinant HBsAg was homogeneous 22-nm particles, suggesting the HBsAg expressed in P. pastoris was self-assembled to virus-like structures. Competitive ELISA indicated that P. pastoris-derived HBsAg possessed the excellent immunoreaction with anti-HBsAg. Animal immunization showed that the immunogenicity of P. pastoris-derived HBsAg was superior to that of S. cerevisiae-derived HBsAg. Together, our results demonstrated that the recombinant HBsAg expressed in P. pastoris could provide promising, inexpensive, and large-scale materials for the diagnostic reagent and vaccine to prevent HBV infection.

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“…Some investigators report that HBV genotypes A‐G can be recognized comparably by commercial HBsAg assays including genotype E [34–37]. In contrast, others found sensitivity differences between HBV genotypes: up to 10‐fold differences in the sensitivity of three commercial assays [38]; lower binding of anti‐HBs by a factor 2–3 to ayw and adr compared to the WHO reference adw [39]; one of 10 HBsAg kits failed to give positive results with genotype E at 0·2 IU/ml [35]; and reduced reactivity in monoclonal antibody binding studies for E/ ayw4 and D/ ayw3 [33]. The results of the 17 highly sensitive HBsAg assays in this study essentially confirm that HBsAg kits used in the EU and Japan detect HBV genotypes A‐F with comparable sensitivity [34,35].…”

Section: Discussionmentioning

confidence: 99%

Performance evaluation of 70 hepatitis B virus (HBV) surface antigen (HBsAg) assays from around the world by a geographically diverse panel with an array of HBV genotypes and HBsAg subtypes

Scheiblauer

El-Nageh²,

Diaz³

et al. 2010

Vox Sanguinis

121

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Background and ObjectivesThis study was conducted by the International Consortium for Blood Safety (ICBS) to identify high-quality test kits for detection of hepatitis B virus (HBV) surface antigen (HBsAg) for the benefit of developing countries.Materials and MethodsThe 70 HBsAg test kits from around the world were evaluated comparatively for their clinical sensitivity, analytical sensitivity, sensitivity to HBV genotypes and HBsAg subtypes, and specificity using 394 (146 clinical, 48 analytical and 200 negative) ICBS Master Panel members of diverse geographical origin comprising the major HBV genotypes A-F and the HBsAg subtypes adw2,4, adr and ayw1-4.ResultsSeventeen HBsAg enzyme immunoassay (EIA) kits had high analytical sensitivity <0·13 IU/ml, showed 100% diagnostic sensitivity, and were even sensitive for the various HBV variants tested. An additional six test kits had high sensitivity (<0·13 IU/ml) but missed HBsAg mutants and/or showed reduced sensitivity to certain HBV genotypes. Twenty HBsAg EIA kits were in the sensitivity range of 0·13–1 IU/ml. The other eight EIAs and the 19 rapid assays had analytical sensitivities of 1 to >4 IU/ml. These assays were falsely negative for 1–4 clinical samples and 17 of these test kits showed genotype dependent sensitivity reduction. Analytical sensitivities for HBsAg of >1 IU/ml significantly reduce the length of the HBsAg positive period which renders them less reliable for detecting HBsAg in asymptomatic HBV infections. Reduced sensitivity for HBsAg with genetic diversity of HBV occurred with genotypes/subtypes D/ayw3, E/ayw4, F/adw4 and by S gene mutants. Specificity of the HBsAg assays was ≥99·5% in 57 test kits and 96·4–99·0% in the remaining test kits.ConclusionDiagnostic efficacy of the evaluated HBsAg test kits differed substantially. Laboratories should therefore be aware of the analytical sensitivity for HBsAg and check for the relevant HBV variants circulating in the relevant population.

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Hepatitis B surface antigen (HBsAg) derived from yeast cells (Hansenula polymorpha) used to establish an influence of antigenic subtype (adw2, adr, ayw3) in measuring the immune response after vaccination

Cited by 31 publications

References 11 publications

Humoral Immune Response Recognizes a Complex Set of Epitopes on Human Papillomavirus Type 6 L1 Capsomers

Humoral Immune Response Recognizes a Complex Set of Epitopes on Human Papillomavirus Type 6 L1 Capsomers

Expression, Purification, and Characterization of Hepatitis B Virus Surface Antigens (HBsAg) in Yeast Pichia Pastoris

Performance evaluation of 70 hepatitis B virus (HBV) surface antigen (HBsAg) assays from around the world by a geographically diverse panel with an array of HBV genotypes and HBsAg subtypes

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