Purification and serology of the W strain of cucumber mosaic virus

During studies on the purification of cucumber mosaic virus (strain W) it was found that preparations were most infective and stable when made from tobacco leaves (10–12 days after inoculation) homogenized in phosphate buffer containing EDTA and thioglycollic acid and clarified with diethyl ether. The preparations were further purified by centrifugation in sucrose density gradients containing EDTA at pH 9.0 and were then stable at 2 °C for > 100 days. When mounted in neutralized ammonium molybdate they were shown to consist of predominantly intact particles. In tube and ring precipitin tests and in agar gel-diffusion tests, specific precipitation with homologous antiserum occurred only in media containing alkaline adjusted solutions (ammonium molybdate and dipotassium hydrogen phosphate).     

Scanning electron microscopy of isolated Chironomus polytene chromosomes

Individual polytene chromosomes have been isolated from Chironomus stigmaterus for scanning electron microscope observations. The three dimensional ultrastructure of these chromosomes consists of a series of chromatin strands extended in the interbands and more tightly coiled or folded in the banded regions. The nucleolus is observed to be a dense disc or doughnut shaped structure surrounding the chromosome while the Balbiani Rings appear as diffuse regions consisting of both fibrillar and granular elements.     

Scanning electron microscopy of isolated peripheral nerve fibres

Summary The surface morphology of normal myelinated nerve fibres prepared in different ways for scanning electron microscopy has been studied and compared with the surface features of similar fibres undergoing retrograde changes. Nodes of Ranvier, paranodal specializations, artefactual fractures of the myelin, and the endoneurial collagen sheaths are described. A regular pattern of elevations, usually with a pitted or depressed surface seen on normal myelinated fibres after certain preparative procedures are thought to be artefacts produced during preparation and to be related to the neurokeratin network.Alterations in the surface structure of fibres central to long-standing nerve transections include irregular protuberances, serial surface corrugations and large swellings, all associated with demyelination. Fibres that have undergone retrograde degeneration consist of endoneurial tubes with focal swellings occupied by macrophages or myelin debris, together with fine unmyelinated and small myelinated regenerating axons. Strict centrifugal progression of myelination of regenerating axons was not observed.We thank Mr. R. A. Willis for his collaboration and for taking the SEM photographs of normal nerve fibres, and the Cambridge Scientific Instrument Co. Ltd. for permission to reproduce the SEM photographs of experimental nerve fibres. We also thank Dr. A. Boyde for access to his SEM and for helpful comments on interpretation of the scanning electron micrographs, Prof. J. Z. Young, Dr. P. K. Thomas, and Dr. R. H. M. King for discussion, and Messrs. P. Reynolds and D. Gunn for photography.A grant from the Muscular Dystrophy Group of Great Britain is gratefully acknowledged.     

Light and electron microscopy of virus inclusions inAmaranthus lividus cells

Summary Amaranthus plants infected with a virus of rod-shaped particles showed under the light microscope intracytoplasmic amorphous and crystalline inclusions.The submicroscopic organization of mesophyll cells from infectedAmaranthus leaves by electron microscopy is described. Besides big crystalline inclusions, long dark inclusions correspondent to needle-like inclusions observed by light microscopy are definable in the cytoplasm. The amorphous inclusion bodies were formed by an overgrown protrusion of vacuolate cytoplasm containing virus particles, long very dark stained inclusions forming dense bands and rings, normal elements of the cytoplasm such as mitochondria, endoplasmic reticulum and ribosomes, and some spherosomes. Inclusions and virus particles were not found in chloroplasts, mitochondria or nuclei of infected cells.     

Purification and properties of peroxidase from Nitrosomonas europaea

Peroxidase from the obligate chemosynthetic bacterium Nitrosomonas europaea was purified 1,500-fold, and its properties were examined. The enzyme had a molecular weight of 53,000 and exhibited characteristic absorption maxima at 410, 524, and 558 mmu. The optimal pH and temperature were 7.5 and 44 C, respectively. The peroxidase reaction had an energy of activation of 5,850 cal/mole and required a primary substrate (H(2)O(2)) concentration of 7 x 10(-6)m to proceed at half maximal velocity (K(m)). Reduced cytochrome, c,p-phenylenediamine and pyrogallol acted as hydrogen donors to the purified peroxidase-H(2)O(2) complex. Conditions most suitable for the chemical oxidation of ammonium by H(2)O(2) were determined. The reaction was rapid and produced nitrite but no nitrate. Hydroxylamine was not detected as an intermediate, but it could substitute for ammonium in the system. Neither the rate nor the extent of these reactions was influenced by purified peroxidase, and no evidence was obtained to support a conclusion that the enzyme performs a vital role in the transformation of ammonium to nitrite by N. europaea.     

High resolution scanning electron microscopy of isolated and in situ cytoskeletal elements

Evidence is presented that cytoskeletal structures (actin filaments, intermediate filaments, and microtubules) can be resolved by scanning electron microscopy after osmium impregnation of biological material, using thiocarbohydrazide as a ligand, followed by critical-point drying. These different classes of filaments or tubules can be identified both as purified protein polymers and as structured organelles within cryofractured or detergent-extracted cells.