Measurement of Cell Wall Volume using Confocal Microscopy and its Application to Studies of Forage Degradation

Abstract

Abstract The distribution of cell wall material between different plant cell types may contribute significantly to the variation in degradability of plant material with a similar overall chemical composition but different anatomy. Assessment of the degradability of cell walls in a section suitable for digestion is a three-dimensional (3-D) problem because of the thickness of section required (50–100 μm). Optical sectioning of thick sections using confocal laser scanning microscopy (CLSM) provides a method of estimating the volume of cell wall material present in tissue sections before and after digestion, and of visualizing the plant tissue using 3-D image reconstruction. The use of CLSM enables degradability measurements to be made on cells in situ and can provide more immediate and relevant information than can be obtained by mechanical fractionation of the tissues. The CLSM method has been used to visualize thick sections taken from maize and barley internodes before and after degradation with cell wall degrading enzymes. Quantitative measurements of cell wall volume and mean cell wall thickness were made on a series of optical sections, and the potential of the method for quantitation of cell wall degradability is assessed.