RESUMO
BACKGROUND AND OBJECTIVES: This 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 METHODS: The 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. RESULTS: Seventeen 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 >or=99.5% in 57 test kits and 96.4-99.0% in the remaining test kits. CONCLUSION: Diagnostic 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.
Assuntos
Antígenos de Superfície da Hepatite B/sangue , Hepatite B/epidemiologia , Técnicas Imunoenzimáticas/métodos , África/epidemiologia , Sequência de Aminoácidos , Ásia/epidemiologia , Países em Desenvolvimento , Reações Falso-Negativas , Genes Virais , Genótipo , Saúde Global , Hepatite B/sangue , Hepatite B/diagnóstico , Hepatite B/virologia , Antígenos de Superfície da Hepatite B/classificação , Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/classificação , Vírus da Hepatite B/genética , Vírus da Hepatite B/imunologia , Humanos , Dados de Sequência Molecular , Kit de Reagentes para Diagnóstico , Sensibilidade e Especificidade , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , América do Sul/epidemiologia , Estados Unidos/epidemiologiaRESUMO
Hepatitis B virus (HBV) genotyping and hepatitis B surface antigen (HBsAg) subtyping were carried out on sera from 196 HBsAg-positive patients, including 151 refugees entering the United States and 45 injection drug users in Seattle. HBsAg subtyping was performed by enzyme immunoassay (EIA) using a panel of monoclonal antibodies and the HBV genotype was determined by polymerase chain reaction (PCR) followed by detection of amplified HBV DNA by a reverse-phase hybridization line probe assay (LiPA) using genotype-specific probes. HBV DNA was detected by PCR in 155 (79%) of the 196 sera and all 155 were genotyped by LiPA. Samples from Southeast Asia were predominantly genotype B/subtype ayw1 and genotype C/adr; samples from the former Soviet Union and eastern Europe were mostly genotype D/ayw2 and genotype D/ayw3; samples from east Africa were mainly genotype A/adw2 and genotype D/ayw2; and samples from injection drug users were mostly genotype D/ayw3 and genotype A/adw2. Some strains of ayw3 gave atypical monoclonal antibody reactivity patterns in the subtyping assay due to a Val/Ala instead of a Thr at amino acid residue 118 and a Thr instead of a Met at residue 125. A strain of ayw2 also gave an atypical monoclonal antibody reactivity pattern due to an Ala instead of a Thr at amino acid residue 123. LiPA genotyping and monoclonal EIA subtyping can provide useful information for epidemiological studies.
Assuntos
Antígenos de Superfície da Hepatite B/análise , Antígenos de Superfície da Hepatite B/imunologia , Antígenos E da Hepatite B/análise , Vírus da Hepatite B/classificação , Vírus da Hepatite B/genética , Hepatite B/virologia , Kit de Reagentes para Diagnóstico , Refugiados , Abuso de Substâncias por Via Intravenosa/virologia , África Oriental/epidemiologia , África Oriental/etnologia , Sequência de Aminoácidos , Anticorpos Monoclonais/classificação , Anticorpos Monoclonais/imunologia , Sudeste Asiático/epidemiologia , Sudeste Asiático/etnologia , Sequência de Bases , Sequência Consenso , Europa Oriental/epidemiologia , Europa Oriental/etnologia , Genótipo , Haiti/epidemiologia , Haiti/etnologia , Hepatite B/epidemiologia , Antígenos de Superfície da Hepatite B/sangue , Antígenos de Superfície da Hepatite B/classificação , Vírus da Hepatite B/imunologia , Humanos , Técnicas Imunoenzimáticas , Pessoa de Meia-Idade , Oriente Médio/epidemiologia , Oriente Médio/etnologia , Noroeste dos Estados Unidos , U.R.S.S./epidemiologia , U.R.S.S./etnologia , Estados UnidosRESUMO
We report the generation of murine triomas by fusing splenocytes from mice previously immunized with HBsAg ay-subtype and a hybridoma, secreting anti-HBsAg ad-subtype monoclonal antibody, which was rendered HGPRT- by induced mutagenesis with N-methyl-N'nitro-N-nitrosoguanidine. The fusion yielded a 83.8% of hybrids showing the antigen specificity of the parental hybridoma and a 16.1% of bi-specific monoclonal antibodies. One of them, coded as 1C8A5, showing a heavy chain isotype (IgG1/IgG2b) was used as capture reagent in an ultramicro-ELISA. As little as 0.78 I.U. of both HBsAg ad- and ay-subtypes could be realiably detected.
Assuntos
Anticorpos Monoclonais/imunologia , Células Produtoras de Anticorpos/imunologia , Anticorpos Anti-Hepatite B/imunologia , Antígenos de Superfície da Hepatite B/imunologia , Hibridomas/imunologia , Imunoglobulina G/imunologia , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/isolamento & purificação , Especificidade de Anticorpos , Fusão Celular , Ensaio de Imunoadsorção Enzimática , Anticorpos Anti-Hepatite B/biossíntese , Anticorpos Anti-Hepatite B/isolamento & purificação , Antígenos de Superfície da Hepatite B/classificação , Humanos , Imunoglobulina G/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C/imunologia , MicroquímicaRESUMO
Foram analisadas 896 amostras de soros HBsAg positivos de cinco regiöes brasileiras. A presença de cinco diferentes subtipos (ayw2, ayw3, ayw4, adw2 e adw4) foi determinada. Na regiäo Norte, os subtipos adw4 (41,2%) e adw2 (37,2%) foram predominantes. No Nordeste somente o subtipo adw2 foi encontrado. No centro Oeste, Sudeste e Sul os subtipos ayw2, ayw3, adw4 estäo presentes, com predominância do adw2 no Centro Oeste e Sudeste (84,3% e 69,4% respectivamente), enquanto que no Sul o subtipo predominante foi ayw3 (41,9%) seguido pelo ayw2 (36,4%). Os subtipos ayw1 e ayr e adr näo foram encontrados entre amostras estudadas. Estes resultados mostram a diferença na incidência dos subtipos do HBsAg em diferentes regiöes do Brasil e seu significado em relaçäo à colonizaçäo e migraçöes neste país
Assuntos
Humanos , Antígenos de Superfície da Hepatite B/classificação , Portador Sadio/sangue , Brasil , Vírus da Hepatite B/imunologiaRESUMO
HBsAg positive serum samples (896) from five brazilian regions were analysed for HBsAg subtypes. The presence of five different subtypes (ayw2, ayw3, ayw4, adw2 and adw4) was detected. In Northern region subtypes adw4 (41.2%) and adw2 (37.2%) were predominant. In the North East only subtype adw2 was encountered. In Central West, South-East and South, subtypes ayw2, ayw3, adw2 and adw4 were present, with predominance of adw2 in Central West and South East (84.3% and 69.4% respectively) whereas in the South the predominant subtype was ayw3 (41.9%) followed by ayw2 (36.4%). Subtypes ayw1, ayr and adr were not found among the samples studied. These results show the difference in the incidence of HBsAg subtypes in the different regions of Brazil and their significance in relation to the colonization and migrations in this country.
Assuntos
Portador Sadio/sangue , Antígenos de Superfície da Hepatite B/classificação , Brasil , Estudos Transversais , Antígenos de Superfície da Hepatite B/análise , Vírus da Hepatite B/imunologia , HumanosRESUMO
A total of 1,915 sera collected in 1979 from asymptomatic hepatitis B surface antigen (HBsAg) carriers were tested for delta antigen, antibody to delta antigen (anti-delta), hepatitis B e antigen (HBeAg) and antibody to hepatitis B e antigen (anti-HBe) in addition to HBsAg and its subtypes. These sera represented blood donated by volunteers to 49 of 57 regions of the American Red Cross located in nine geographic regions of the United States and Puerto Rico. A total of 72 (3.8%) sera had anti-delta activity while none had a detectable level of delta antigen. A significantly higher (p less than 0.01) prevalence of anti-delta (12.1%) was found in San Jose, California (Pacific Region); on the other hand, the East South Central region covering Alabama, Kentucky, Mississippi and Tennessee had a significantly lower (p less than 0.05) prevalence (1.4%) of anti-delta when compared with all other regions combined. Anti-delta was, however, detected in all regions of the United States and in Puerto Rico. The cause of significant differences in the prevalence of anti-delta was not clear. The distribution of anti-delta was not associated with age, sex or blood type of the donor. Sixty-nine of 70 samples with anti-delta were found among the 1,527 samples that had either HBeAg or anti-HBe. And among 149 that lacked both HBeAg and anti-HBe, only one sample had anti-delta. The difference is statistically significant (p less than 0.05). The presence of anti-delta was not associated with HBsAg/ad (2.7%) or HBsAg/ay (4.6%).