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At least 73 records · Page 4Linked to original sources
PubMed1990

Cold stage design for high resolution electron microscopy of biological materials.

Both the number and range of applications of cryotechniques in transmission electron microscopy are increasing rapidly. In some cases, most notably the determination of protein structure by electron crystallography, progress has been limited by the performance of commercially available cryo stages. We review the design and performance criteria for stages which will be necessary for wide applicability in high resolution studies of biological specimens. The important criteria include an operating temperature below -140 degrees C with a low rate of contamination of the specimen, ability to tilt to 60 degrees, and perhaps most important, good resolution as judged by an effective modulation transfer function of 0.8 at 0.35 nm. Most applications also require an effective cryotransfer system. Up until now, most work in high resolution electron crystallography has been accomplished with laboratory-built stages which meet some, but not all, of these criteria. The availability of cold stages which fully meet criteria will allow the rapid expansion of high resolution studies by electron microscopy in structural biology.

Cryopreservation↗
PubMed1997

High resolution imaging of native biological sample surfaces using scanning probe microscopy.

The possibility of acquiring high resolution topographs using scanning probe microscopes under physiological conditions allows the observation of biomolecules at work. Progress has recently been made in imaging protein-DNA complexes, individual oligomers and protein arrays. Scanning probe microscopes are now tools that complement X-ray crystallography and electron microscopy.

Bacterial Outer Membrane Proteins↗
PubMed1989

Ultrahigh resolution scanning electron microscopy of biological materials.

In recent years, several ultrahigh resolution scanning electron microscopes (SEM) were successively developed. They were all equipped with a field emission electron gun and an objective lens with a short focal length, and showed a resolution better than 1 nm. With such instrument, not only intracellular structures but also virus, bacteriophages, and biological macromolecules were clearly observed. With improvement of the instrumental resolution, some unexpected problems came to the fore in the area of specimen preparation. The first is the metal coating of the specimen, because coated metal particles are plainly seen as rounded "pebbles" at very high magnifications. For observations at the high magnifications, therefore, uncoated and not conductively stained specimens were used. The second problem is contamination by the electron beam. This problem is complicated and remains unsolved. Although the ultra-high resolution scanning electron microscopy has just begun, it will surely open new research fields in biomedicine.

Animals↗
PubMed1992

Ultrahigh-resolution scanning electron microscopy of biological materials.

The age of ultrahigh-resolution scanning electron microscopy (SEM) began in 1985, when the UHS-T1, with a resolution of 0.5 nm, was developed. Commercial instruments of the same or similar types followed rapidly. As instrumental resolution progressed, conventional specimen preparation methods became inadequate, and a number of new techniques were devised. In this paper, detailed procedures for these preparation methods such as the CC plate method and heavy metal impregnation are described, together with precautions recommended for achieving ultrahigh-resolution. Some applications of the method to biological specimens are also reported. Morphological identification of immunoglobulins prepared from human blood was attempted, and although the identification was not completely successful this technique may yet come to be of use in the clinical examination of allergic or infectious diseases. SEM images of complement, Clq, proteoglycan and the helical structure of double stranded DNA are shown, as also is the visualization of immunolabelled cell-surface receptors.

Carbon↗
PubMed1991

Application of high-resolution scanning electron microscopy to biological macromolecules.

The development of ultrahigh-resolution scanning electron microscopes (SEMs) has made the observation of biological macromolecules feasible, but adequate preparation methods have not yet been established. Although it has been possible to observe some molecules after they have been spread on a carbon substrate, this method has not proved suitable for other molecules which exhibit lower contrast, or are more susceptible to damage by the electron beam. In this study we have applied heavy-metal impregnation methods using phosphotungstic acid, uranyl acetate, or osmium tetroxide mordanted by tannic acid. In addition, contamination due to the electron beam was reduced by improving the vacuum in the specimen chamber, and by the use of a heated specimen stage. Using these measures, haemocyanin, ferritin, apoferritin, thyroglobulin and immunoglobulin M were successfully image. Ultrahigh-resolution SEM seems likely to become an important means for studying the morphology of biological macromolecules.

Animals↗
PubMed1995

Cryo atomic force microscopy: a new approach for biological imaging at high resolution.

A low-temperature atomic force microscope (cryo-AFM), operated in liquid nitrogen vapor, has been constructed for biological applications. The system provides an adjustable imaging temperature from 77 to 220 K with atomic resolution achieved on crystalline specimens. Imaging with NaCl microcrystals demonstrates that the system is free from surface contamination. Below 100 K, several biological specimens, including immunoglobulins and DNA as well as red blood cell ghosts, were imaged at high spatial resolution. Measurements on individual macromolecules showed that the mechanical strength is significantly greater at cryogenic temperatures with an estimated Young's modulus 1000-10,000 times that of a hydrated protein at room temperature, providing a solid basis for future improvements and applications of cryo-AFM in structural biology.

DNA↗
PubMed1980

Hereditary genius: a centennial problem in resolution of cultural and biological inheritance.

Galton believed that his data on hereditary genius support purely genetic inheritance. On the contrary, analysis under a simple model shows no clear resolution of genetic and cultural inheritance in this material, or generally, unless adoptive relations, monozygous twins, and/or environmental indices are included in a design adequate to test the assumptions under which they are informative.

Adult↗
PubMed1988

Biological microanalysis by secondary ion mass spectrometry: status and prospects.

Secondary ion mass spectrometric (SIMS) analysis of biological problems is an evolving technique. Lateral resolution of currently available commercial instrumentation estimated from actual samples is 0.5 micron, and subcellular organelles can be distinguished. The interrelationship of lateral resolution, elemental concentration and ionizability are, however, important in controlling the actual lateral resolution achievable. Although depth resolutions of 5 nm have been measured in other systems, no test of depth resolution in biological systems has been done, and this parameter is also concentration and ionization dependent. The development of liquid metal ion sources in combination with scanning ion microprobes has a potential lateral resolution of as little as 20 nm, but initial studies with this instrumentation show that tissue preservation at the submicron level becomes an important issue. The current development of a cold-transfer stage for SIMS instruments may obviate the problem of submicron localization of diffusible elements, and initial studies indicate that much more needs to be understood about the ionization process in hydrated samples. Quantitation of diffusible elements using external standards has been achieved over a 30 micron diameter analyzed area. Strategies for analysis of areas limited to 1 micron or less has been suggested using image processing techniques, which take advantage of the lateral resolution inherent in the ion optical system. Matrix effects in biological tissues have been reported and constitute a serious problem for analysis of biologicals which must be addressed for each question. However, development of laser ionization of sputtered particles may both increase the sensitivity of analysis and decrease the importance of ionizability of elements. Chemical analysis of organic molecules is another use of SIMS, but, at present, at the cost of losing localized information. SIMS analysis of biological samples is being systematically evaluated and requires increased accessibility of this instrumentation to the end-user for full development of its role in physiological problems.

Elements↗
PubMed1983

Resolution and reconstitution of biological pathways from 1919 to 1984.

The central theme of this paper is the reconstitution of the Warburg effect, the high aerobic glycolysis of malignant tumors. The history of resolution-reconstitution started with the isolation of glycolytic enzymes. In 1945 Meyerhof prepared an extract from yeast that did not ferment unless an ATPase was added. An extract of Ehrlich ascites tumor cells that does not glycolyze in the presence of catalytic amounts of Pi and nucleotides without addition of an ATPase is presented as a model for future reconstitutions of the Warburg effect. Natural polypeptide preparations from placenta and hypothalamus 1) stimulate a protein kinase from tumor plasma membranes, 2) serve as substrates for another protein kinase from tumor plasma membranes, and 3) stimulate glycolysis of normal rat or chick embryo fibroblasts and may be related to the transforming tumor growth factors. We can hope that an exploration of the mechanism by which these polypeptides stimulate glycolysis could lead to the successful reconstitution of the Warburg effect in an in vitro system. It may also help us understand how tumor RNA or malignant DNA induces the various other biochemical changes that take place when normal cells are transformed to tumor cells.

Animals↗
PubMed2001

Enantiomers of 2-[(Acylamino)ethyl]-1,4-benzodiazepines, potent ligands of kappa-opioid receptor: chiral chromatographic resolution, configurational assignment and biological activity.

Compounds 2a and 3a-e are racemic 2-[(acylamino)ethyl]-1,4-benzodiazepines, tifluadom analogs, with high affinity and selectivity towards the kappa-opioid receptor. We describe the enantiomeric separation of all compounds through liquid chromatography with chiral stationary phases, as well as the resolution of the enantiomers of the most interesting compounds, 2a and 3a, by the semipreparative column Chiralpak AD. The configuration of the resolved enantiomers was investigated: the comparative study of CD and (1)H NMR spectra shows that compounds (-)-2a and (-)-3a have the same absolute configuration of (+)-(S)-tifluadom. A study on the stereoselective interaction with opiate receptors is reported.

Benzodiazepinones↗
PubMed1978

Radiation damage in the high resolution electron microscopy of biological materials: a review.

Radiation damage to a biological specimen arises from a variety of interactions between the illuminating electrons and the atoms in it. The relative probabilities of these events, and the amout of energy transferred, can be calculated from basic physical theory. The microscopic damage caused in a particular specimen in given operating conditions is more difficult to predict, but it can be measured by a number of macroscopic indicators, the chief of which are loss of mass and changes in the energy loss spectrum (or electron diffraction, pattern, if any). For most biological material the observed rate of damage is such as to set a limit to the intensity of illumination, the maximum magnification and the minimum size of detail that can be made visible. Several techniques have been devised and tested for reducing the radiation sensitivity of a specimen, of which cooling to a very low temperature and encasing it in an inert medium are the most effective. If the various protective measures act cooperatively, they could increase the effective resolution of sensitive material by an order of magnitude, making possible electron microscopy of the atomic structure of, for instance, the nucleic acid bases and other macromolecules. The prospects for observing living cells at a resolution better than that of the best optical microscopes would remain very small.

Biological Products↗
PubMed1994

High-resolution electron microscopy of biological specimens in cubic ice.

Images of two biological test specimens, catalase and TMV, were recorded in cubic ice, prepared by controlled devitrification at -130 degrees C and -75 degrees C or in vitrified buffer. Cubic ice provides a rigid support for biological specimens which is stable under the electron beam to within about 1 A, as shown by images of the ice lattice. Neither catalase nor TMV were disrupted by the crystallization of vitrified water. Electron diffraction patterns of highly oriented rafts of TMV extending to 2.3 A resolution were used to judge the quality of TMV images. Structural high-resolution details of TMV was better in cubic ice, and the success rate for recording good images was higher than in vitrified medium.

Catalase↗
PubMed1976

Resolution of cultural and biological inheritance by path analysis.

Analysis of family resemblance is developed in terms of three genetic parameters, six parameters for cultural inheritance, and one parameter for an index estimating family environment. With efficient use of nuclear families the model is fully determinate. Other biological and social relationship provide additional degrees of freedom for testing goodness of fit. Performance of the model is satisfactory on simulated data with extreme gene-environment interaction. Applied to a large body of published data on I.Q., neither genetic assortative mating nor gene-environment covariance is significant by a likelihood ratio test, but heritability is less and cultural inheritance is greater for adults than children. Whereas family resemblance of children is largely genetic, for adults it is largely due to their childhood environments, presumably acting on occupational aspirations. Further resolution is more likely to come from nuclear families than from the rare relationships that were favored by classical human genetics.

Culture↗
PubMed1988

3-dimensional imaging of biological structures by high resolution confocal scanning laser microscopy.

Imaging in confocal microscopy is characterized by the ability to make a selective image of just one plane inside a specimen, virtually unaffected -within certain limits- by the out-of-focus regions above and below it. This property, called optical sectioning, is accompanied by improved imaging transverse to the optical axis. We have coupled a confocal microscope to a computer system, making the combination of both an excellent instrument for mapping the 3-dimensional structure of extended specimens into a computer memory/data array. We measured that the volume element contributing to each data point has, under typical fluorescence conditions, a size of 0.2 X 0.2 X 0.72 micron. The data can be analysed and represented in various ways, i.e., stereoscopical views from any desired angle. After a description of the experimental arrangement, we show various examples of biological and food-structural studies. The microscope can be operated either in reflection or in fluorescence. In the latter mode a spectral element allows selection of the wavelength band of fluorescence light contributing to the image. In this way, we can distinguish various structures inside the cell and study their 3-dimensional relationships. Various applications in biology and the study of food structure are presented.

Animals↗
PubMed1999

Resolution, molecular structure and biological activities of the D- and L-enantiomers of potent anti-implantation agent, DL-2-[4-(2-piperidinoethoxy)phenyl]-3-phenyl-2H-1-benzopyran.

Compound 1 (DL-2-[4-(2-piperidinoethoxy)phenyl]-3-phenyl-2H-1-benzopyran, CDRI 85/287) a potent anti-estrogen and anti-implantation agent has been successfully resolved into its pure D- and L-enantiomers. Biological studies showed L-enantiomer to be the active form, exhibiting a fivefold higher receptor affinity for the rat uterine cytosolic estrogen receptor, 100% contraceptive efficacy at 1.3 mg/kg dose in single day schedule and 89% inhibition of estradiol induced increase of uterine weight at its contraceptive dose. The absolute stereochemistry determined by X-ray crystallographic analysis showed that the L-enantiomer has 2R configuration at its asymmetric centre.

Animals↗