Detection of Vesicular-Arbuscular Mycorrhizal Fungus Colonization of Roots by Using a Dot-Immunoblot Assay

Abstract

An immunoblotting procedure was developed to overcome the difficulty in identifying root colonization by a vesicular-arbuscular mycorrhizal fungus. The procedure utilized a murine monoclonal antibody that reacts with a protein in spores and hyphae of Glomus occultum , a fungus characterized by abundant production of hyaline spores and nonstaining intraradical infection. Minimally disturbed whole roots were squashed on nitrocellulose membranes. After inactivation of endogenous peroxidase, an indirect enzyme-linked immunosorbent assay was performed on the nitrocellulose with peroxidase-conjugated anti-mouse antibody as the second antibody. Antigen from G. occultum , revealed by a precipitating stain, was seen as purple dots on the nitrocellulose, which also retained the impression of the root.