In Situ Localization of Citrus tristeza virus Inclusion Bodies with Antibodies Developed to the Recombinant p20 Non-Structural Protein

摘要

An in situ immunoassay is described for localization of Citrus tristeza virus (CTV) inclusion bodies by light microscopy with antibodies developed to the recombinant p20 non-structural protein. Free hand bark sections of ca. 100 mum tick from healthyor CTV infected plants were fixed with 70 ethanol and subsequently incubated with specific polyclonal antiserum anti-p20 either from rat or rabbit at dilutions 1:5,000 y 1:10,000 y 1:1,000 and 1:3,000 respectively and IgG anti species at 1:30,000 dilution; at the end, the chromogenic solution NBT/BCIP was added. Viral inclusion bodies were visualized by light microscopy at 10 and 40X magnifications as dark blue irregular areas in phloem associated tissues in CTV infected plants; no dark blue color wasobserved in the phloem tissues of healthy plants.