Biomedical subjects

D A Grano

Publications and source records attributed to D A Grano.

3 recordsLinked to original sources

PubMed1983

Quantitative evaluation of high-resolution features in images of negatively stained Tobacco Mosaic Virus.

This study investigates the causes of the apparent differences between the optical diffraction pattern of a micrograph of a Tobacco Mosaic Virus (TMV) particle, the optical diffraction pattern of a ten-fold photographically averaged image, and the computed diffraction pattern of the original micrograph. Peak intensities along the layer lines in the transform of the averaged image appear to be quite unlike those in the diffraction pattern of the original micrograph, and the diffraction intensities for the averaged image extend to unexpectedly high resolution. A carefully controlled, quantitative comparison reveals, however, that the optical diffraction pattern of the original micrograph and that of the ten-fold averaged image are essentially equivalent. Using computer-based image processing, we discovered that the peak intensities on the 6th layer line have values very similar in magnitude to the neighboring noise, in contrast to what was expected from the optical diffraction pattern of the original micrograph. This discrepancy was resolved by recording a series of optical diffraction patterns when the original micrograph was immersed in oil. These patterns revealed the presence of a substantial phase grating effect, which exaggerated the peak intensities on the 6th layer line, causing an erroneous impression that the high resolution features possessed a good signal-to-noise ratio. This study thus reveals some pitfalls and misleading results that can be encountered when using optical diffraction patterns to evaluate image quality.

Computers↗

PubMed1981

Periodic surface array in Caulobacter crescentus: fine structure and chemical analysis.

A periodic array structure on the cell surface of Caulobacter crescentus CB15 was revealed by electron microscopy of the cell envelope, using negative staining, thin-sectioning, and freeze-etching. This structural layer has been isolated from liquid cultures, in which large pieces of the two-dimensional array are shed by cells grown to high density. Often areas of intact array corresponding to the entire cell surface could be found. The hexagonally arranged structure was highly ordered and had an unusual degree of complexity, as determined by optical diffraction and computer processing of micrographs of negatively stained, isolated surface array. Filtered, reconstructed images were obtained from both normal and low-electron-dose micrographs demonstrating resolutions of 2.9 and 25 nm, respectively. Comparison by optical diffraction and image filtering of micrographs recorded by using either normal or minimal beam exposure techniques suggested that the lower-resolution features of the image are very stable to electron exposure. Gel electrophoresis indicated that isolated array preparations contain a number of polypeptides. It appears likely that more than one of these proteins are structural components of the array, in contrast to a single protein found in many bacterial surface arrays. The Caulobacter surface array is also unusual in that the repeated units are widely spaced with no apparent direct connection. Computer spatial averaging provided information about the shape and complexity of the connecting elements, and this was compared with some additional electron microscopic evidence of linking structures. Thin-sectioning studies confirmed the image features seen by other techniques, but the addition of tannic acid in the fixation procedure was required to visualize the structure. A comparison of these results with out current knowledge of the Caulobacter cell envelope suggests interesting questions about the biogenesis of this membrane structure and its involvement in the cell development process of this organism.

Bacteria↗

PubMed1978

Molecular orientation of bacteriorhodopsin within the purple membrane of Halobacterium halobium.

The direction of orientation of the protein bacteriorhodopsin within the purple membrane of Halobacterium halobium has been determined by selected-area electron diffraction of membranes preferentially oriented by adsorption to polylysine. Purple membrane is known to adsorb preferentially to polylysine by its cytoplasmic surface at neutral pH and by its extracellular surface at low pH. To maintain the adsorbed membranes in a well-ordered state in the electron microscope, an improved technique of preparing frozen specimens was developed. Large areas of frozen-hydrated specimens, devoid of bulk water, were obtainable after the specimen was passed through a Ca stearate film at an air-water interface. High-resolution microscopy was used to relate the orientation observed in the electron diffraction patterns to the orientation of the projected structure that is obtained from images. We have found that the three-dimensional structure determined by Henderson and and Unwin [Henderson, R. & Unwin, P.N.T. (1975) Nature 257, 28--32] is oriented with the cytoplasmic side uppermost--i.e., the helices fan outward on the cytoplasmic side of the membrane.

Bacteriorhodopsins↗