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Detection of double-stranded RNA by ELISA and dot immunobinding assay using an antiserum to synthetic polynucleotides

1991; Elsevier BV; Volume: 33; Issue: 1-2 Linguagem: Inglês

10.1016/0166-0934(91)90002-h

1879-0984

J. Aramburu, Jesús Navas‐Castillo, Pedro Moreno, M. Cambra,

Plant Pathogenic Bacteria Studies

An antiserum against polyinosinic-polycytidylic acid (In-Cn) was used to detect double-stranded RNA (dsRNA) by several serological techniques. DsRNA was readily detected by indirect ELISA (ELISA-I) and dot immunobinding assay (DIA). Addition of the antigen to poly-L-lysine-precoated plates and blocking with uncreamed milk powder allowed detection levels of 100 pg.ml-1 In-Cn by ELISA-I. Concentrations as low as 1 ng.ml-1 were detected by DIA using polyvinyliden difluoride (PVDF) membranes. Detection capacity with nitrocellulose membranes was 1000 times lower than with PVDF. ELISA-I and DIA enabled detection of dsRNA in enriched fractions from cucumber mosaic virus (CMV)- and citrus tristeza virus (CTV)-infected plants and from virus-infected Penicillium chrysogenum mycelium. These techniques showed similar or higher sensitivity for detection of dsRNA than separation by polyacrylamide gel electrophoresis and silver staining.