Isolation of Cuticle and Vascular Bundles of Leaves by Maceration in Rumen Fluid

We report a new macerating technique for plant leaves which permits the isolation of cuticle and vascular bundles. The maceration medium is rumen fluid, a complex mixture of interacting microorganisms, used full strength or diluted with a nutrient buffer solution in the ratio of 2:5. An incubation period of up to 72 hours at 39 C permits cuticle and vascular-bundle networks to be isolated. The technique is illustrated with fresh leaf samples from pearl millet, Pennisetum typhoides; orchardgrass, Dactylis glomerata; and alfalfa, Medicago sativa.