Resin-casting: a method for investigating apoplastic spaces

A method has been developed in which liquid resin can be injected or infiltrated into spaces within a plant body, for example the lumens of vessels, fibers, and intercellular spaces. After polymerization of the resin, plant tissues are digested away with two solutions used in sequence: first, equal parts concentrated hydrogen peroxide and glacial acetic acid; second, concentrated sulfuric acid. Complete digestion renders three-dimensional casts of the spaces in the original tissue. These can be examined with scanning electron microscopy, light microscopy, or a dissecting microscope. Casts have such high fidelity and high resolution that details of pit canals, pit chambers, and perforation plates can be studied. Vessel casts over 15 cm long and revealing the details of several thousand constituent vessel elements have been obtained easily. Casts of the lumens of all cell types and of narrow intercellular spaces are obtained by prolonged infiltration with a low-viscosity resin solution. Alternatively, rapid, brief injection of the resin by vacuum produces casts predominantly of just those spaces that were open to the resin at the cut ends of the sample, for example the lumens of vessels and secretory ducts or the intercellular space network of an aerenchymatous parenchyma.