Método imunoenzimático usando antígeno delta derivado do soro para detecção sérica de antígeno e anticorpo do vírus da hepatite delta

One of the main difficulties related to the detection of the Hepatitis Delta Virus (HDV) antigen and antibody has been the source of the needed HD antigen since HDV containing human and animal livers are very difficult to obtain and since yield is low. This fact prompted us to try to use the serum of patients in the acute phase of HDV infection as a source of HDAg and turn to enzyme immunoassays (EIA) instead of RIA for the sake of easiness and economy in the amount of HDAg needed. The antigen for EIA was obtained from patients during the acute phase of HDV infection and the antibody from patients who have been carriers for many years. For the detection of the antigen, a sandwich type method was employed, whereas for the antibody a competition assay was developed. In order to assess the relative specificity and sensibility of the test, the antibody assay was compared to a commercial RIA (C. RIA, Abbott) and to a non-commercial RIA (NC RIA). Forty-two sera were tested by the two methods and only in two cases discrepant results were obtained. Its is concluded that: 1) sera from patients in the acute and chronic phases of HDV infection can be used as source of both antigen and antibody, for immunoassays; 2) EIA and RIA have comparable relative specificity and sensibility and 3) EIA is easier to perform, cheaper, non-hazardous, has a longer shelf-life and saves scarce HDAg.Um dos maiores problemas no desenvolvimento de metodologia para detecção de antígenos e anticorpos do vírus da hepatite delta (VHD) tem sido a fonte de antígenos, uma vez que fígados humanos e de animais infectados pelo VHD são de difícil obtenção e baixo rendimento Ao uso de soro de pacientes na fase aguda da infecção pelo VHD, como fonte de antígenos, associamos a técnica imunoenzimática, com finalidade de facilitar o manuseio e economizar antígeno. O antígeno delta foi obtido a partir do soro de indivíduo na fase aguda de infecção por VHD e o anticorpo a partir de soro de portadores crônicos de VHD. Para a detecção do antígeno foi empregado o método "sanduíche" e para a detecção do anticorpo um ensaio tipo competição. Visando testar a especificidade e a sensibilidade relativas do novo método de detecção de anticorpo, foram feitas comparações do mesmo com radio-imunoensaio comercial (C-RIE, Lab. Abbott) e um radio-imunoensaio desenvolvido na Unité 271-INSERM de Lyon, França (NC-RIA). Soros de 42 pacientes foram testados pelos três métodos, sendo observados resultados discrepantes em apenas 2 casos. Os autores concluem que: 1) soros de pacientes na fase aguda e crônica da infecção pelo VHD podem ser usados como fonte de antígeno e anticorpos em ensaios imunológicos 2) o EIE e o RIE tem especificidade e sensibilidade relativas comparáveis 3) o EIE é de fácil execução, mais barato, não poluente, tem vida útil maior, além de proporcionar economia de antígeno

An ELISA method using serum derived HDAg for the sorological detection of HDV antigens and antibodies

One of the main difficulties related to the detection of the Hepatitis Delta Virus (HDV) antigen and antibody has been the source of the needed HD antigen since HDV containing human and animal livers are very difficult to obtain and since yield is low. This fact prompted us to try to use the serum of patients in the acute phase of HDV infection as a source of HDAg and turn to enzyme immunoassays (EIA) instead of RIA for the sake of easiness and economy in the amount of HDAg needed. The antigen for EIA was obtained from patients during the acute phase of HDV infection and the antibody from patients who have been carriers for many years. For the detection of the antigen, a sandwich type method was employed, whereas for the antibody a competition assay was developed. In order to assess the relative specificity and sensibility of the test, the antibody assay was compared to a commercial RIA (C. RIA, Abbott) and to a non-commercial RIA (NC RIA). Forty-two sera were tested by the two methods and only in two cases discrepant results were obtained. Its is concluded that: 1) sera from patients in the acute and chronic phases of HDV infection can be used as source of both antigen and antibody, for immunoassays; 2) EIA and RIA have comparable relative specificity and sensibility and 3) EIA is easier to perform, cheaper, non-hazardous, has a longer shelf-life and saves scarce HDAg