Resolution Of Light Microscopy Research Articles

An electron-microscopic study of the early outgrowth from chick spinal cord in vitro.

1. A simple method is described for the location and ultrathin sectioning of selected areas of the outgrowth from chick spinal cord in vitro. 2. What appear by light microscopy to be single outgrowing fibres are commonly (but not invariably) bundles of processes. Some are so small as to be beyond the limits of light microscopical resolution. Over appreciable distances bundles may have no supporting cells associated with them. 3. Within the outgrowth, synaptic profiles have been observed, and it is suggested that they are newly formed during culture.

The Fine Structure and Histochemistry of the Infective Eggs of Dipylidium caninum

The ultrastructure and histochemical composition of the infective eggs of Dipylidium caninum were studied. The uterine capsule is a cellular modification of the uterine lining. Quantities of acid mucopolysaccharide-positive vitelline material within the uterus and uterine capsule form a layer at the surface of the outer capsule of the egg. This material cements the outer capsules together to form clusters of eggs. In the outer capsule a thin lamina separates the outer material from an inner homogeneous PAS-positive layer. Histochemically, a structural polysaccharide or glycoprotein composition is indicated in the outer capsule. The cytoplasmic layer, beneath the outer capsule, contains cell fragments, lipid droplets, mitochondria, and alpha-glycogen. The embryophore, which surrounds the hexacanth embryo, is composed of two layers of rods at right angles to each other. The rods elicit a positive reaction for keratin. The oncosphere contains germinative and somatic cells, six keratinaceous hooks with associated embryonic musculature, and a penetration gland. The latter, in the mature oncosphere, has a granular cytoplasm containing small dense elongate bodies. In the infective oncosphere, the cytoplasm of the penetration gland is modified and the dense bodies become swollen and lose their internal structure. The morphology of the infective eggs of Dipylidium caninum (Cyclophyllidea; Dilepididae) has been studied with light microscopy. Observations on this and related dilepidid species (Ogren, 1955, 1958, 1959a, 1959b, 1961, 1962; Rybicka, 1961, 1964, 1966a, 1966b; Silverman, 1954) have not elucidated completely the morphology and composition of structures seen in this stage. Johri (1957) undertook histochemical studies on the eggs of a related cyclophyllidean family, the Taeniidae. No such studies appear for the Dilepididae. The embryophore and its formation in the taeniid egg have been studied with electron microscopy by Morseth (1965). This investigation and a single electron micrograph published in a study by Race et al. (1966) are the only works on the fine structure of cyclophyllidean eggs. Much of the structure of the cyclophyllidean egg is just beyond the resolution of light microscopy, and there is little information available concerning the composition of egg structures. This investigation, utilizing elecReceived for publication 19 May 1967. * A thesis submitted to the Louisiana State University Medical Center Graduate School as part of the requirements for the degree of Master of Science. This investigation was partially supported by Public Health Service Research Grant AI-02347 from the NIAID of the NIH. tron microscopic and histochemical techniques on the mature egg of Dipylidium caninum, was undertaken to gain information in these areas. MATERIALS AND METHODS Adult Dipylidium caninum were obtained at autopsy from the intestine of naturally infected dogs. Worms were removed rapidly and the terminal gravid proglottids dissected from the strobila. For election microscopy: Gravid proglottids, containing eggs, were cut into small pieces and fixed at 4 C in 3.82% glutaraldehyde buffered at pH 7.4 (Pease, 1964). After 1 to 8 hr in the fixative, the tissue was washed in cold Millonig's buffer, and postfixed 1 to 2 hr in cold 2% buffered osmium tetroxide (Millonig, 1961). The tissue was washed again in Millonig's buffer, rapidly dehydrated in graded concentrations of ethyl alcohol beginning with 50%, and passed to a 1:1 mixture of propylene oxide and Araldite 502 overnight. Embedment was in Araldite 502 according to the method of Luft (1961). Sections were cut with glass knives on a PorterBlum MT-I microtome and mounted on naked 200to 400-mesh copper grids. These were stained 10 min in 1.5% uranyl acetate (Watson, 1958), followed by lead citrate (Reynolds, 1963), with brief intermediate and final washings in carbonate-free glass-distilled water. Preparations were examined with an RCA EMU 2B electron microscope operated at 50 kv. Initial magnifications were from 1,900 X to 18,700 X. For histochemistry: Whole gravid proglottids were fixed in 10% formalin or in glutaraldehyde at the same concentration used for electron microscopy. Tissue was washed in tap water for several hours, dehydrated in graded concentrations of

Electron Microscopy of Leaf Surfaces

THE morphology of leaf surfaces is of considerable interest in the study of the effects of selective herbicides. In the course of investigation into the nature of some leaf surfaces, it was suspected that structure beyond the resolution of the light microscope was present. The electron microscope was therefore employed in an attempt to determine the nature of this structure. The examination of the surface of a leaf required the use of a replica technique, preferably not involving the treatment of the specimen with organic solvents, since many leaf surfaces are covered with a waxy layer which might be affected. A single-stage method involving evaporated material was therefore indicated, and though at first it seemed likely that any such method would meet with considerable difficulties, an extremely interesting technique was easily developed.