Life Sciences Commons^™

Combined Bronchoalveolar-Vascular Casting Of The Canine Lung, John A. Nettum

Scanning Microscopy

Canine bronchoalveolar and vascular corrosion casts were prepared using unfixed tissue and Dow-Corning Room Temperature Vulcanizing Silastic® 734. The casts were observed using stereo light microscopy and scanning electron microscopy. The casts show the relationship between the vasculature and airway and demonstrate intricate microanatomical details. Also, microvasculature filling was enhanced using unfixed tissue as compared to my previously described technique using fixed tissue. Prewashing the microvasculature with cold phosphate buffered saline appeared to facilitate microvascular filling with silicone rubber. The described method is useful for rapidly making durable models for studying the normal respiratory airway and microvasculature. It should be …

Application Of Monte Carlo Simulations To Critical Dimension Metrology In A Scanning Electron Microscope, Jeremiah R. Lowney

Scanning Microscopy

The state of the art in Monte Carlo simulations of scanning electron microscope (SEM) signals is reviewed. Two Monte Carlo computer codes were written to simulate the transmitted-, backscattered-, and secondary-electron signals from targets in a scanning electron microscope. The first, MONSEL-11, is applied to semi-infinite lines produced lithographically on multi-layer substrates. The second, MONSEL-111, is an extension to fully three-dimensional targets. Results for a 1 μm step, etched in a silicon substrate are compared with experimental data. The comparisons show that it is possible to obtain edge locations to an uncertainty of less than 10 nm. Simulations were performed …

Atomic Force Microscopy And Optical Studies Of Organic Thin Films With Hydrogen-Bonded Networks, Arkadi L. Litvin, Valery N. Bliznyuk, Vladimir V. Tsukruk, Suresh Valiyaveettil, David L. Kaplan

Scanning Microscopy

Brewster angle microscopy and atomic force microscopy were used to characterize the surface morphology of thin films in situ or after transfer onto solid supports. Two acids were studied, differing in carboxylic acid head groups, resulting in significantly different morphological features for thin films formed from these two amphiphiles on a Langmuir trough. Differences in self-assembly and domain sizes were correlated with the formation of hydrogen-bonded networks. The influence of surface hydrophobicity or hydrophilicity during deposition on morphology was also characterized, with spherulitic features appearing in some samples.

Monte Carlo Simulation Of Electron Backscattering In Solids Using A General-Purpose Computer Code, E. Acosta, E. Coleoni, G. Castellano, J. A. Riveros, J. M. Fernández-Varea, F. Salvat

Scanning Microscopy

A Monte Carlo study of backscattering of kilovolt electrons in solids, a process of primary importance in electron microscopy and surface analytical techniques, is carried out. Simulations have been performed using the general-purpose simulation code PENELOPE (an acronym for "Penetration and ENErgy LOss of Positrons and Electrons"), which generates electron-photon showers in arbitrary materials. A systematic comparison of results from PENELOPE with available experimental data, and with results from simulations with a much more sophisticated code, is given for electron beams with energies between 2.5 and 60 keV and elemental solids with atomic numbers Z = 4 to 92. It …

Effects Of Castration Upon The Morphology Of The Accessory Sex Organs Of The Male Rat - A Scanning Electron Microscopy Study, R. Wahlqvist, E. Dahl, K. J. Tveter

Scanning Microscopy

A systematic, comparative study of the accessory sex glands of the adult male rat after androgen withdrawal was carried out. The changes were investigated by using scanning electron microscopy at different intervals after surgical castration. The main common signs of epithelial cell involution were flattening of the cell surface, reduction of the size and number of microvilli, some blurring of the cell borders, cessation of secretory activity and diminution of the luminal volume of the glands. Overall, confident signs of atrophy were evident after one week, and complete epithelial involution was reached by the third week. The epithelial cell atrophy …

Emerging Applications Of Fluorescence Spectroscopy To Cellular Imaging: Lifetime Imaging, Metal-Ligand Probes, Multi-Photon Excitation And Light Quenching, Joseph R. Lakowicz

Scanning Microscopy

Advances in time-resolved fluorescence spectroscopy can be applied to cellular imaging. Fluorescence lifetime imaging microscopy (FLIM) creates image contrast based on the decay time of sensing probes at each point in a two-dimensional image. FLIM allows imaging of Ca²⁺ and other ions without the need for wavelength-ratiometric probes. Ca²⁺ imaging can be performed by FLIM with visible wavelength excitation. Instrumentation for FLIM is potentially simple enough to be present in most research laboratories. Applications of fluorescence are often limited by the lack of suitable fluorophores. New, highly photostable probes allow off-gating of the prompt autofluorescence, and measurement of …

Scanning Electron Microscopy Of The Accessory Sex Glands Of The Adult Male Rat, R. Wahlqvist, E. Dahl, K. J. Tveter

Scanning Microscopy

This study describes the morphology of the accessory sex glands of the adult male rat as observed by scanning electron microscopy (SEM). The purpose was to obtain a systematic and comparative SEM description of these glands and to evaluate different preparation techniques. A common morphological feature is polyhedral delineation of the cells, which exhibited a variable convexity of their apical surface. The cell apices were more or less studded with microvilli. Nevertheless, it was possible to distinguish the glands by their surface morphology. In the ventral prostate, there was a considerable heterogeneity in cell surface appearance. The lateral lobe bad …

Hydrogen Atom Extraction And Redeposition On Hydrogen-Terminated Silicon Surface With Scanning Tunneling Microscope At Room Temperature, D. H. Huang, Y. Yamamoto

Scanning Microscopy

We have previously demonstrated that hydrogen atoms on the hydrogen-terminated Si(100)-2xl:H surface can be extracted by applying either positive or negative voltage pulses between an STM tip and the sample surface with a certain tunneling current. This phenomenon suggests that electron excitation and field evaporation both play a role in atom transfer. Under our experimental conditions, hydrogen atoms are extracted in pairs from either a single dimer or from two adjacent dimers in neighboring rows. Hydrogen atoms can also be redeposited onto the Si(100)-2xl:H surface from a tungsten tip, which has picked up hydrogen from the sample surface through the …

The Role Of Calcium In The Regulation Of Apoptosis, David J. Mcconkey

Scanning Microscopy

The recognition that apoptosis is regulated by an evolutionarily conserved set of polypeptides from the nematode Caenorhabditis elegans to humans suggests that a conserved set of biochemical mechanism(s) may also be involved in the response. Early evidence suggested that the endogenous endonuclease implicated in apoptosis in most model systems is Ca2+ -dependent, and subsequent work from a number of independent laboratories suggests that alterations in cytosolic Ca2+ homeostasis are one of the conserved biochemical pathways regulating the response. Molecular targets for Ca2+ are now being identified and include signal transduction intermediates, endonuclease(s) and proteases, and the enzymes involved in the …

Influence Of Bleeding On Post-Mortem Tenderization Of Fish Muscle During Chilled Storage, M. Ando, A. Nishiyabu, T. Nakagawa, Y. Makinodan

Scanning Microscopy

The influence of bleeding on post-mortem tenderization of fish muscle during storage was studied. Six fish species were used for the present study. Among them, delay of post-mortem tenderization of muscle by bleeding was shown in the pelagic fishes: yellowtail, horse mackerel, and striped jack. On the other hand, bleeding had no influence on the muscle firmness of the bottom fishes: red sea bream, flatfish, and rudder-fish. According to transmission electron microscopy, degradation of pericellular collagen fibrils was delayed in bled yellowtail and horse mackerel. In the case of striped jack, though, collagen fibril degradation could not be observed; slower …

Applications Of Laser Microprobe Mass Spectrometry In Biology And Medicine, Annette Bakker, Luc Van Vaeck, Wim Jacob

Scanning Microscopy

Laser microprobe mass spectrometry (LMMS) provides spot analysis with a lateral resolution of 1-5 μm. Focused laser ionisation, combined with mass spectrometry, yields information on elements, molecular identification of inorganic substances, and structural characterisation of organic molecules of typically less than 2-3 kDa. Quantification in complex heterogeneous systems, such as biological materials, is not possible at the present state of the art. The strength of the method lies in the qualitative information on the molecular composition of the analyte, not just element detection, with a lateral resolution at the light microscopy level. The applications of this technique in the field …

Contact Force Dependence On Relative Humidity: Investigations Using Atomic Force Microscopy, E. Finot, E. Lesniewska, J. -C. Mutin, S. I. Hosain, J. -P. Goudonnet

Scanning Microscopy

This paper deals with the ability of scanning force microscopy to determine contact forces of various materials. Indeed, with high spring constants at low relative humidity, the nature of the material can be determined by measurement of the contact force as the tip approaches. Cantilevers with a high spring constant are used to achieve solid-solid contact for the tip-sample system. The capillary force estimation provides information on the development of the height of the water meniscus formed between the tip and different surfaces depending on the relative humidity. Finally, we focus our attention on measurements of moduli of elasticity which …

Covalent Labeling Of Proteins With Fluorescent Compounds For Imaging Applications, Darl R. Swartz

Scanning Microscopy

The labeling of proteins with fluorescent compounds for microscopy has allowed a greater understanding of biological processes. The preparation of fluorescent proteins is the first step in development of their use in microscopy. Methods are described to label and characterize a protein as an example of the general approach for other proteins. Skeletal muscle alpha-actinin was labeled with either fluorescein-5-maleimide or 5-iodoaceamidofluorescein and the reaction characterized. The maleimide reaction was much more rapid and efficient than the iodoacetamide reaction giving a coupling efficiency of 65% under the given ration conditions. The fluorescein-5-maleimide alpha-actinin was functionally characterized and there was essentially …

Labeling With Nanogold And Undecagold: Techniques And Results, James F. Hainfeld

Scanning Microscopy

A significant new development in gold labeling for microscopy has been achieved through the use of gold cluster compounds that are covalently attached to antibodies or other probe molecules. These unique gold probes are smaller than most colloidal gold conjugates and exhibit improved penetration into tissues, higher labeling densities, and allow many new probes to be made with peptides, nucleic acids, lipids, drugs, and other molecules. A new fluorescent-gold conjugate is useful for examining localization by fluorescence microscopy, then visualizing the same label at the ultrastructural level in the electron microscope.

Quantitative Analyses Of Microwear And Honing On The Sloping Crest Of The P3 In Female Japanese Monkeys (Macaca Fuscata), Teruyuki Hojo

Scanning Microscopy

Female wild Japanese monkeys (Macaca fuscata), as with all male cercopithecoids, use the mesiobuccal surfaces or the elongated crests of the mandibular third premolars (P3s), as cutting blocks that wear against edges of maxillary canines during threat manifestation or food-eating. In other words, the crests of their P3s are honed with the maxillary canines. The crests become sloped during growth and more heavily striated with the advance of age. The number, directions, lengths, and widths of these striations have been analyzed quantitatively using scanning electron microscopy (SEM). Two samples showed two distinct types of parallel striations, one longer and thicker …

The Neonatal Chinchilla Cochlea: Morphological And Functional Study, R. V. Harrison, J. R. Cullen, S. Takeno, R. J. Mount

Scanning Microscopy

The developmental time scale of the cochlea varies from species to species. We investigate here the condition of the neonatal cochlea in the chinchilla, a species increasingly used in auditory research. We have examined the morphology of cochlear hair cells using scanning microscopy, and the development of auditory function during the first postnatal month by monitoring auditory brainstem evoked responses (ABR). We find that although there were some outer hair cell kinocilia present in middle and apical areas, the hair cells otherwise were mature at 24 hours after birth. Furthermore, cochlear auditory thresholds are adult-like at birth. However, whilst there …

Advanced Instrumentation And Methodology Related To Cryoultramicrotomy: A Review, H. Sitte

Scanning Microscopy

This review is concerned with the considerable progress in the field of cryo-ultramicrotomy (cryofixation, cryosectioning, investigation and analysis of cryosections) during recent years. This progress includes both more efficient instrumentation and methodology. The article is mainly directed to the investigation and analysis of frozen-hydrated sections in the low dose cryo-transmission electron microscopy (TEM) and cryo-energy filtered TEM (EFTEM). A general survey is followed by an evaluation of the different relevant procedures. Both cryo-ultramicrotomy for macromolecular cytochemistry (Tokuyasu technique) and cryo-ultramicrotomy for element analysis are only shortly mentioned without discussion of the chemical and analytical approach. Because of lack of first …

Cytoskeletal Changes During Radiation-Induced Neoplastic Transformation Of Human Prostate Epithelial Cells, S. C. Prasad, P. J. Thraves, J. S. Rhim, M. R. Kuettel

Scanning Microscopy

We recently reported tumorigenic transformation of SV 40-immortalized neonatal human prostate epithelial cells (267Bl) by exposure to fractionated doses of X-rays. Altered morphology and anchorage independence were observed following two successive fractions of 2 Gy each (F3-SAC). Additional 2 Gy treatments to these non-tumorigenic cells to a total dose of 30 Gy resulted in radiation-transformed tumorigenic colonies (267Bl-SXR). Malignant transformation of parental 267B 1 cells was also achieved by consecutive 2 Gy exposures to a total dose of 30 Gy (267Bl-XR). This study discusses the cytoskeletal changes in the F3-SAC, 267Bl-XR and 267Bl-SXR derivatives of these human prostate epithelial cells. …

Pre-Embedding Staining Of Single Muscle Fibers For Light And Electron Microscopy Studies Of Subcellular Organization, Evelyn Ralston, Thorkil Ploug

Scanning Microscopy

Skeletal muscle fibers are large, multinucleated cells which pose a challenge to the morphologist. In the course of studies of the distribution of the glucose transporter GLUT4, in muscle, we have compared different preparative procedures, for both light (LM) and electron microscopy (EM) immunocytochemistry. Here we show that pre-embedding staining of single teased fibers, or of single enzymatically dissociated fibers, has several advantages over the use of sections for observing discrete patterns that extend over long distances in the cells. We report on an optimization study carried out to establish fixation and permeabilization conditions for EM immunogold labeling of the …

Structural And Calcification Patterns Of The Neonatal Line In The Enamel Of Human Deciduous Teeth, Tetsuo Kodaka, Tsuneyoshi Sano, Shohei Higashi

Scanning Microscopy

The neonatal line of the enamel in human deciduous teeth was observed by scanning electron microscopy using secondary and backscattered electron signals. The neonatal line containing irregular structures of enamel prisms with disordered crystal arrangements was basically formed by the abrupt bending of the prisms towards the root. Usually, the prisms gradually bent back again to regain their previous orientation, but the prisms in the inner and the surface layer sometimes ran straight ahead after bending. The prism sheath regions showing hypocalcification contained a relatively large amount of organic material as shown by treatment with chromium sulfate. When etched with …

Monte Carlo Simulation Of Secondary Electron Emission From Thin Film/Substrate Targets, K. Murata, M. Yasuda, H. Kawata

Scanning Microscopy

We have developed a Monte Carlo simulation model of secondary electron emission from thin film/substrate samples, taking into consideration their exact boundary condition. First, the validity of the model is checked in comparison with the experimental data reported such as the secondary electron emission and backscattering yields from thick Al, thick Au targets and Al thin films on a Au substrate, the energy distribution of secondary electrons, and the contribution of backscattering to the secondary electron emission yield. The agreement is relatively good. Next, we have applied the model to the secondary electron emission from Au films on an Al …

Generation Of High Efficiency Ssdna Hybridization Probes By Linear Polymerase Chain Reaction (Lpcr), Gregory W. Konat

Scanning Microscopy

The polymerase chain reaction (PCR) methodology can be employed to produce DNA hybridization probes. The major advantages of this paradigm over other techniques include superior specific activity of the probes, the versatility of sequence selection, the ability to produce short probes, and the simplicity of the procedure. We have further improved the efficiency of PCR probes by generating single stranded (ssDNA) probes that do not reanneal with themselves in solution, and hence, their availability for the interaction with the complementary sequences of the target is profoundly increased. Protocols for ³²P-dCTP labeled and digoxigenin-dUTP labeled probes have been elaborated to …

Microhardness And Scanning Electron Microscopy Analysis Of Nd:Yag Laser And Acid Treatment Effects In Dentin, J. M. White, G. L. Adams

Scanning Microscopy

Laser irradiation at and above parameters producing the modification threshold for dentin causes structural changes to the dentin surface. This study determined the microhardness of dentin before and after laser modification and acid treatment using a repeated-measures de-sign. Seven dentin sections (4 mm in thickness) were cut from freshly extracted non-carious third molars using a diamond saw. The middle occlusal third was used as the test dentin surface. One section served as a control (C); three received laser modification (L) and then acid treatment (L + A); and three received acid (A) and then laser treatment (A + L). Laser …

Experimental Nephrolithiasis In Rats: The Effect Of Ethylene Glycol And Vitamin D3 On The Induction Of Renal Calcium Oxalate Crystals, R. De Water, E. R. Boevé, P. P. M. C. Van Miert, G. Deng, L. C. Cao, Th. Stijnen, W. C. De Bruijn, F. H. Schröder

Scanning Microscopy

Using ethylene glycol (EG) and vitamin D3 as crystal-inducing diet (CID) in rats, we investigated the effect of the dosage of EG on the generation of chronic calcium oxalate (CaOx) nephrolithiasis. We collected weekly 24 hour urines and measured herein the amount of oxalate, calcium, glycosaminoglycans (GAG's), creatinine, protein, alkaline phosphatase (AP), 𝝲-glutamyl transpeptidase (𝝲GT), and N-acetyl-𝝱-glucosaminidase (NAG). The potential of these urines to inhibit crystal growth and agglomeration was also evaluated. After four weeks, the kidneys were screened by histology and radiography for the presence of CaOx crystals and the amount of kidney-associated oxalate was biochemically measured. Using 0.5 …

Problems In Preparation Of Chromosomes For Scanning Electron Microscopy To Reveal Morphology And To Permit Immunocytochemistry Of Sensitive Antigens, A. T. Sumner

Scanning Microscopy

Although much information about chromosome structure and behaviour has been obtained using light microscopy, greater resolution is needed for a thorough understanding of chromosome organisation. Scanning electron microscopy (SEM) can provide valuable data about these three-dimensional organelles. The introduction of methods using osmium impregnation of methanol-acetic acid-fixed chromosome spreads revolutionised matters, producing life-like images of chromosomes. Nevertheless, it became clear that osmium impregnation introduced various artefacts, although the resulting images were still useful. Methanol-acetic acid-fixed chromosomes are, in fact, flattened on the glass substratum, and the 3-dimensional appearance obtained after osmium impregnation is the result of swelling during this process. …

Preparation Of Plasmid Dna In Transfection Complexes For Fluorescence And Electron Spectroscopic Imaging, Marek Malecki

Scanning Microscopy

The aim of this project was to develop procedures necessary to study mechanisms of receptor mediated gene transfer by means of integrated microscopy.

Plasmid DNA was incorporated into a transfection complex consisting of poly(L)lysine and transferrin to which the nuclear localization signal was conjugated. This complex was presented to cultured glioma cells. Preparation of the transfected DNA for imaging was pursued by two methods. In the first method tetramethylrhodamine, nanogold, and ferritin were linked through streptavidin to the biotinylated plasmid DNA. Trafficking of the fluorescent derivatives was studied in living cells with fluorescence microscopy. Then, selected cells were rapidly cryo-immobilized. …

Common Epidermal Lesions Of The Bowhead Whale, Balaena Mysticetus, William G. Henk, Dianna L. Mullan

Scanning Microscopy

Samples of common skin abnormalities from 23 subsistence-harvested bowhead whales (Balaena mysticetus) were examined. Most lesions fell into three broad classes: shallow lacerations, circular depressions, and epidermal sloughing. Both circular depression lesions and epidermal sloughing lesions may be divided into more than one subgroup based on morphological criteria. Examination of each of the subgroups using light and scanning electron microscopy suggests relationships among the subgroups of a class. These proposed relationships are discussed, as are some possible etiologies. Scanning electron microscopy reveals abundant bacteria and diatoms present in association with each lesion class but no evidence of a particular association …

Adhesion And Endocytosis Of Calcium Oxalate Crystals On Renal Tubular Cells, Y. Kohjimoto, S. Ebisuno, M. Tamura, T. Ohkawa

Scanning Microscopy

The present investigation was designed to study interactions between Madin-Darby canine kidney (MOCK) cells and calcium oxalate monohydrate (COM) crystals and to clarify the significance of these crystal-cell interactions in stone pathogenesis.

MOCK cells cultured in the presence of COM crystals showed a time-dependent uptake of crystals; this was specific for COM crystals. In the dynamic model system designed to study these phenomena under more physiological conditions, COM crystals adhered to the cell surface and were subsequently internalized. In this endocytotic process, the microvilli of the cell appeared to play an important role. The observation by scanning electron microscopy of …

X-Ray Microscopy: Preparations For Studies Of Frozen Hydrated Specimens, A. Osanna, C. Jacobsen, A. Kalinovsky, J. Kirz, J. Maser, S. Wang

Scanning Microscopy

X-ray microscopes provide higher resolution than visible light microscopes. Wet, biological materials with a water thickness of up to about 10 μm can be imaged with good contrast using soft X-rays with wavelengths between the oxygen and carbon absorption edges (at 24 and 43 Å). The Stony Brook group has developed and operates a scanning transmission X-ray microscope (STXM) at the National Synchrotron Light Source (NSLS) at Brookhaven National Laboratory. The microscope is used for imaging with a current resolution of 50 nm, and for elemental and chemical state mapping.

Radiation damage imposes a significant limitation upon high resolution X-ray …

Low Calcium Diet In Hypercalciuric Calcium Nephrolithiasis: First Do No Harm, Bernhard Hess

Scanning Microscopy

Many studies indicate that up-regulated production of 1,25(OH)2-vitamin D3 (calcitriol) with increased intestinal absorption of calcium is the primary event causing idiopathic hypercalciuria. Thus, a low calcium diet appears to be a straightforward strategy in calcium stone formers with hypercalciuria (HCSF). However, the efficacy of such a regimen has never been established, and lowering calcium intake from 1000 to 400 mg/day further enhances calcitriol production. On a diet chronically restricted in calcium, many stone formers increase their intake of animal flesh protein. The latter is known to increase renal mass, and calcitriol levels indeed are positively correlated with renal mass …