Anticuerpos monoclonales contra proteínas de la nucleocápside del virus IBR y su evaluación por ELISA
Monoclonal antibodies against proteins of the IBR virus nucleocapside and their assessing by ELISA
Keywords:
herpesvirus bovino-I, protefnas virales, inmunología, glicoprotefnas, nucleocapside (es)bovine herpesvirus-I, viral proteins, immunology, glycoproteins, nucleocapside (en)
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A partir de un aislamiento de una cepa de campo del virus de IBR, se produjeron clonos productores de anticuerpos monoclonales (AcMo)contra componentes de su nucleocápside, para 10 cuallos inóculos para ser descargados en los ratones Balb/c fueron obtenidos del cultivo de la línea celular (MDBK), los cuales se purificaron y concentraron por centrifugaci6n en gradiente continuo de sacarosa al 30% y posterior precipitación por métodos químicos. Las nucleocápsides obtenidas fueron sometidas a SDS-PAGE preparativas al 100%. Se empleó la técnica de ELISA directa como prueba de tamizaje para los sobrenadantes de los hibridomas positivos para la producción de los AcMo. Se obtuvieron ocho hibridomas positivos al virus de IBR por ELISA; cinco fueron positivos por prueba de seroneutralización y por inmunodot; realizada la selección clonal por medio del proceso de dilución límite, se obtuvieron 56 clones positivos al virus de IBR, que reconocen la proteína de 39.8kDa; sus sobrenadantes fueron confirmados por las pruebas de ELISA, Dot y Western Blot. Con los AcMo, se normalizó una técnica de ELISA de captura, que permite detectar tanto la cepa de campo como la cepa de referencia Iowa.
Monoclonal antibodies were produced against the nucleocapside of a field IBR virus strain. The virus was growth in MOBK cell line. Viral components were purified and concentrated in a continuous 30% sacarose gradient followed by chemical precipitation. Nucleocapsides were run in a 10% SDS-PAGE gel. Positive hybridomes were tested using a direct ELISA developed in our laboratory. As a result eigth ELISA positive clones were obtained, from these five were also positive to seronetralization and immunodot. The clones recognize a 39.8Kda protein. Aditionally, a capture-ELISA was developed using the monoclonal antobodies from this research. This ELISA is useful to detect a reference as a field nm virus strain.
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Copyright (c) 2002 Revista Colombiana de Biotecnología
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