RESUMO
Hepatitis B virus (HBV) is one of the main pathogens responsible for hepatitis and hepatocellular carcinoma. Human plasma-derived Hepatitis B immune globulin (HBIG) is being used for prophylactic and liver transplantation currently. However, it may be necessary to replace a HBIG with a recombinant one because of limited availability of human plasma with high anti-HBsAg antibody titer and possible contamination of human pathogens. A Chinese hamster ovary (CHO) cell line, HB-C7A, was established which produces a fully human IgG1 that binds HBsAg. The HB-C7A exhibits approximately 2600 units/mg of antibody. The affinity (K(a)) of HB-C7A is 1.1 x 10(8) M(-1) by Biacore analysis and estimated 6.7-fold higher than that of Hepabig (a plasma-derived HBIG from Green Cross Corp., Yongin, Korea) by competition ELISA. The HB-C7A recognizes the conformational "a" determinant of HBsAg and binds HBV particle more efficiently than the Hepabig. The HB-C7A binds to HBV-infected human liver tissue but does not bind to normal human tissues. This HB-C7A has several advantages compared to plasma-derived Hepabig such as activity, safety and availability.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Superfície da Hepatite B/imunologia , Hepatite B/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/genética , Afinidade de Anticorpos/imunologia , Reações Antígeno-Anticorpo/imunologia , Células CHO , Cricetinae , Cricetulus , Reações Cruzadas/imunologia , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Mapeamento de Epitopos , Hepatite B/virologia , Vírus da Hepatite B/imunologia , Vírus da Hepatite B/isolamento & purificação , Humanos , Imunoglobulina G/biossíntese , Imunoglobulina G/genética , Imunoglobulina G/imunologia , Imuno-Histoquímica , Imunoprecipitação , Fígado/imunologia , Fígado/virologia , Dados de Sequência Molecular , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/isolamento & purificaçãoRESUMO
A monoclonal antibody (Mab) against ochratoxin A (OTA) was produced from the hybridoma cell line C7G25, which was established by the fusion of Sp2/0-Ag14 myeloma cells with spleen cells isolated from a BALB/c mouse immunized with the OTA-bovine serum albumin conjugate. This Mab belongs to the IgG(2a) heavy-chain subclass with a kappa-type light chain. The level of 50% inhibition concentration was 1.20 ng/mL in a competitive direct enzyme-linked immunosorbent assay (cdELISA), and the detection limit was 0.12 ng/mL. This antibody is specific for OTA but also shows cross-reactivity with ochratoxin B (31.7%) in a cdELISA. On the basis of the sandwich format using the produced Mab against OTA, a rapid immunochromatographic assay was developed to efficiently detect OTA. This method was able to detect up to 500 ng/mL of OTA in <10 min.