Abstract
A dot enzyme linked immunosorbent assay (dot-ELISA) was previously developed to detect specific antibodies in rabbits sera immunized against FIA protein obtained from Yersina pestis. This antigen was covalently linked onto the surface of dacron (polyethyleneterephthalate). Here, standard conditions are described for the optimization of this procedure: an amount of 20 ng of FIA protein was fixed onto dacron; anti-rabbit IgG peroxidase conjugate diluted 1:8,000 and 30% non-fat instant milk as blocking substance were used throughout the method. This procedure was compared with that employing nitrocellulose as solid-phase which showed to be more sensitive. However, the method based on dacron did not show false positive reactions against non-immunized rabbits sera at low antigen amount and diluted anti-IgG peroxidase conjugate.
dot-ELISA; dacron; Yersinia pestis; solid-phase
Standardization of the dot enzyme-lynked immunosorbent assay (dot-ELISA) for experimental plague
Silvia M. L. Montenegro1
Alzira M. P. de Almeida1
Luiz B. Carvalho Júnior2
FIOCRUZ, Centro de Pesquisas Aggeu Magalhães, Recife, Brasil
UFPE, Departamento de Bioquimica, Laboratório de Enzimologia, Recife, Brasil
A dot enzyme linked immunosorbent assay (dot-ELISA) was previously developed to detect specific antibodies in rabbits sera immunized against FIA protein obtained from Yersina pestis. This antigen was covalently linked onto the surface of dacron (polyethyleneterephthalate). Here, standard conditions are described for the optimization of this procedure: an amount of 20 ng of FIA protein was fixed onto dacron; anti-rabbit IgG peroxidase conjugate diluted 1:8,000 and 30% non-fat instant milk as blocking substance were used throughout the method. This procedure was compared with that employing nitrocellulose as solid-phase which showed to be more sensitive. However, the method based on dacron did not show false positive reactions against non-immunized rabbits sera at low antigen amount and diluted anti-IgG peroxidase conjugate.
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Publication Dates
-
Publication in this collection
01 June 2009 -
Date of issue
Mar 1993
