VIEW ARTICLE

Techniques

Enzyme-Linked Immunosorbent Assay for Plant Viruses in Intact Leaf Tissue Disks. C. Peter Romaine, Assistant professor, Department of Plant Pathology, The Pennsylvania State University, University Park 16802; Susan R. Newhart(2), and David Anzola(3). (2)(3)Research graduate students, Department of Plant Pathology, The Pennsylvania State University, University Park 16802. Phytopathology 71:308-312. Accepted for publication 23 July 1980. Copyright 1981 The American Phytopathological Society. DOI: 10.1094/Phyto-71-308.

Tobacco ringspot virus (TRSV) and maize dwarf mosaic virus (MDMV) were detected by enzyme-linked immunosorbent assay (ELISA) when leaf tissue disks were substituted for leaf extracts as the test sample. The procedure entailed incubating 6-mm-diameter disks of leaf tissue, sampled with a paper punch, in microtiter plate wells containing 200 μ1 of phosphate-buffered saline with Tween-20 and polyvinyl pyrrolidone. When a single virus-infected leaf disk was assayed, the absorbance value (A_{405 nm}) obtained was poorly reproducible, ranging from 0 to 50% of that observed by testing the leaf extract. However, a more comparable test reliability was obtained by increasing the 2-hr substrate reaction time to 24 hr and by incubating several leaf disks in each plate well. By using this protocol, MDMV in corn and TRSV in geranium were detected from 1 wk up to at least 2 mo postinoculation with a reliability comparable to the conventional test using a leaf extract. The leaf disk sampling technique, when used to complement the ELISA test, should prove valuable for the rapid and sensitive qualitative evaluation of virus infection when dealing with large numbers of plants.