A METHOD FOR DETERMINING PHYTOALEXIN CONCENTRATIONS IN FLUORESCENT, HYPERSENSITIVELY NECROTIC CELLS IN COTTON LEAVES

Average phytoalexin concentration in the hypersensitively necrotic cells of a leaf sample was computed by dividing the sample's phytoalexin content, determined by extraction and HPLC, by the water content of those mesophyll cells that exhibited yellow-green fluorescence and/or brown pigmentation. Mesophyll water content was determined by measuring total tissue water content and multiplying it by the fraction of the tissue water in the palisade and spongy mesophyll, determined by stereological analysis of cross sections. The percentage of the mesophyll cells that were yellow-green fluorescent and/or brown was determined by counting such cells, as well as total cells, in 24 microscope fields per sample. The error (coefficient of variation of the mean) in these determinations of phytoalexin concentration was estimated by propagation of error analysis to be ±20%. This strategy for determining cellular phytoalexin concentrations is applicable to other host/pathogen systems for which there is a means of determining which cells contain phytoalexin. © 1992.