Keizo Maruyama

Stereoscopic scanning electron microscopy of the chromosomes in Vicia faba (broad beans)

Metaphase chromosomes of Vicia faba (broad beans) were observed in situ with a scanning electron microscope by cryofracturing the cell. The chromosome is composed of tortuous fibers 500 A in diameter. They may be seen distributed randomly, but sometimes they are seen running parallel in the chromosome. The parallel fibers spiral around the main body of the chromosome, whereas they run longitudinally in the secondary constriction. If a chromosome is composed of a single strand (unineme hypothesis for the chromosome), the parallel fiber arrangement may imply that a single fiber is looping back and forth in the chromosome.

Localization of fluorescent substances in the cellular slime moldDictyostelium discoideum cells during growth and development

The localization of fluorescent substance was observed microscopically in livingDictyostelium discoideum cells. The fluorescence was localized in the vacuoles of the vegetative cells. The fluorescent vacuoles were not observed in the dead cells. The fluorescent vacuoles in the cytoplasm were lost in starved cells which are able to form an aggregate and to differentiate. The fluorescent vacuoles were not lost but decreased slightly in the cytoplasm of full grown cells and of cells grown in liquid nutrient medium for an extended period of time (stationary phase cells). On a solid substratum, fluorescent vacuoles were also lost from the cells, where the vegetative cells aggregate and form a slug-shaped mass of cells. The whole slug showed homogeneous fluorescence. In a finally constructed fruiting body, the spore mass showed fluorescence. In a spore mass, the fluorescence was not observed in the spores but in the interspore space of the spore mass. It is suggested that vegetative cells secrete fluorescent substance into the inter-cellular space in the mass of cells during development.

Growth pattern of isolated zoospores in Hydrodictyon reticulatum (Chlorococcales, Chlorophyceae)

Zoospores at various developmental stages in Hydrodictyon reticulatum were isolated from parent cells and cultured in Waris medium. Isolated zoospores grew to mature vegetative cells, and were able to reproduce zoo‐spores that formed daughter hexagonal nets. Three types of shape appeared in cells 24 h after isolation: cylindrical, Y‐shaped and 4‐armed type. Protrusions of Y‐shaped or 4‐armed cells were formed at an angle of about 120° to the long axis of the cell. When cells were isolated at later stages, more cells became cylindrical in shape and fewer ceils became Y‐shaped or 4‐armed, Direction of cell growth also seemed to depend largely on the developmental stages of the zoospores. The later the isolated stages were, the more the cells elongated along the long axis of the zoospores.

Lumazine-like Fluorescence in a Mass of Spores of the Cellular Slime Mold, Dictyostelium discoideum

Using a fluorospectrophotometer, we examined the fluorescence of a crude preparation from the spore masses ofDictyostelium discoideum. Fluorescence emission spectra and excitation spectra suggested that the fluorescence of the crude preparation was a lumazine-like fluorescence rather than a pterin-like fluorescence. By using a microspectrophotometer, we observedin situ the fluorescence emission of a lumazine-like substance localized only in the spore mass of the fruiting body.

INTRACELLULAR RIBONUCLEASE ISOZYME ACTIVITY OF THE CELLULAR SLIME MOLD CELLS, DICTYOSTELIUM DISCOIDEUM AX2 AND DICTYOSTELIUM MUCOROIDES

The isozymes of ribonuclease were analyzed in cell‐free, crude extracts of cells by reversed activity staining of polyacrylamide gradient gels after electrophoresis. We previously reported three isozymes of the intracellular ribonuclease in Dictyostelium discoideum NC4, which were found in vegetative cells. In the present study, we examined the ribonucleolytic activity of an axenic strain of D. discoideum Ax2 cells which can grow with or without bacteria. The mobility of these isozymes (Dd I, II and III) was the same regardless of the difference in the feeding condition. We also found the three isozymes of D. mucoroides cells, Dm I, II and III. Dm I was different from Dd I in D. discoideum cells, while Dm II and Dm III were the same as Dd II and Dd III in terms of the mobility in polyacrylamide gradient gels during electrophoresis.

Dilute aldehyde treatment on aldehyde-fixed cells for observing chromosomes in situ with the scanning electron microscope

A new preparation method is introduced to reveal intracellular structures in the scanning electron microscope and its application to mitotic cells in root meristems of Vicia faba is demonstrated. The root tips are fixed with a mixture of formaldehyde and glutaraldehyde and the fixed tissues are frozen and fractured in liquid nitrogen. They are then incubated successively in dilute solutions of aldehyde (formaldehyde or glutaraldehyde) and osmium tetroxide. By this treatment, the excess cell‐matrix is removed from the fractured surface of the cell, and a deep view into the cell‐interior can be obtained with the scanning electron microscope. Varied levels of substructure are observed on the surface of chromosomes.

IMMUNOGLOBULINS OF LYMPH NODES

Present-day concepts of antibody-globulins are predicated upon the existence of several different kinds of y-globulin (immunoglobulins), which are named as IgG (yJ, IgM (=yIM, BzM), IgA (=yIA, FzA), etc. on the basis of their immunochemical characters. Several studies have been published on which type of cells concern the synthesis of each immnnoglobulin, however, there is some difference in opinion concerning cytological relationship between plasma cells and other immunocompeterit cells. At least, IgG and IgM are easily detectable by immunoeleotrophoresis in rabbits. In the present study, rabbit lymph node cells engaged in IgG and IgM synthesis have been stained by the fluorescent technique to determine which type of cells is involved with the respective formation of immunoglobulins. Fluorescent antisera employed in the experiments are specific for heavy chains of IgG and IgM which are class specific antigens. It turned out that the plasma cells are mainly concerned with IgG and large lymphocytes including lymphogonia with IgM.