bluuebunny/crossref_metadata_2025 huggingface dataset:
Recent Developments in Mechanical Specimen Preparation for Tem and Sem Applications
Authors: Li W , Q. Wang S , Trussell R , Xu M , Venables RD , Venables D , Maher D , Hirsh EA
Published: July 2020

The continued reduction in the size of critical features in integrated circuits has resulted in the need to develop rapid, site-specific, sectioning techniques to enable efficient physical characterization of the structures of interest. We have implemented a mechanical polishing approach to achieve this objective with the additional goals of maximizing the number of targeted sites in a sample that can be analyzed, and minimizing physically destructive procedures, such as ion beam exposure. Precision sample preparation approaches have been under investigation for both transmission electron microscopy and scanning electron microscopy.

The mechanical specimen preparation approach used in this work is a variant of the well-known wedge polishing technique. Here we use a polishing tool that does not contact the grinding surface, thus allowing precise control of the wedge angle. Prior to sample preparation, the polishing tool head was precision aligned parallel to the platen.
Development And Applications Of Highly Precise Phase Measurement Technique Using Phase-Shifting Electron Holography
Authors: Yamamoto K , Kawajiri I , Tanji T , Hibino M , Hirayama T
Published: July 2020

Today's information-oriented society requires high-density, high-quality magnetic recording media. For the development of such new recording materials, the quantitative observation of magnetic fine structures by electron holography is eagerly awaited. However, the magnetic fields around particles smaller than 50 nm have not been observed because they are too weak to be observed in the usual way. Here, we report a highly precise phase measurement technique, improved phase-shifting electron holography. Using this method, we observed weak electric and magnetic fields precisely. The precision of the reconstructed phase image was as good as 2π/300 rad.

In phase-shifting electron holography, the phase images are reconstructed from a series of electron holograms whose interference fringes are shifted relative to one after another. The shifting of interference fringes is achieved by tilting the incident electron beam, which corresponds to shifting the initial phase of the incident electron waves in a specimen plane.
Live Electron Holography: A Window to the Phase World
Authors: Voelkl E
Published: July 2020

While electron holography has been around for many years (1948, see [1]), the process of obtaining a view into the phase world stayed a tedious one for a long time. When CCD cameras became available and started to replace film, many attempts were made to obtain a “live” view into the phase world. By far the most successful setup for a live phase display is described in [2]. There, Chen et al. use a mixture of digital and analog techniques to obtain phase images at TV-rate. This setup allows to move the sample and/or watch time dependent specimen changes in a unique way, i.e., live in the phase world. The reason why Chen uses a mixture between analog and digital methods is the capability of an optical lens system for fast Fourier processing.

Computers have become significantly faster since Chen's introduction of the mixed setup in 1992 and allow to process holograms rapidly and obtain about one phase image per second on present standard computer systems and acquisition cameras.
Scanning Force Microscopy and Nanomangulation: Studies of Dna and Proteins Involved in Dna Repair
Authors: Erie Dorothy , Ratcliff Glenn , Guthold Martin , Bullock Valerie , Pliske Michelle , Superfine Richard , Taylor Russell
Published: July 2020

Repair of damaged or incorrectly matched DNA is essential to the survival of all organisms. Consequently cells have devised a plentitude of pathways for repair. We have been investigating the mechanisms of mismatch repair and base excision repair. Both of these repair processes involve a large number of proteins that interact with one another as well as with DNA. Our long-term goal is to assemble complexes that are fully functional for DNA repair and to image the process of DNA repair. In addition, we wish to i) determine the stoicheometry of binding of the protein complexes to each other and to DNA, ii) monitor conformational changes due to substrate binding, iii) measure physical properties of DNA and the complexes. To accomplish this end, we have endeavored to improve techniques for solution imaging as well as those for data analysis. In this presentation I will discuss data on the stoicheometry of binding in several protein complexes and data on the physical properties of DNA.

To measure the physical properties of DNA, we utilize a nanoManipulator, a modified Scanning Force Microscope with a novel, user-friendly interface that allows easy and controlled manipulation of nanometer-sized samples.
Structural Forces in Biomolecules
Authors: Rief Matthias , Oesterhelt Filipp , Clausen-Schaumann Hauke , Gaub Hermann E
Published: July 2020

The high spatial resolution and the force sensitivity of the Atomic Force Microscope have made it possible to perform mechanical experiments with single molecules. These experiments give direct access to the forces that stabilize biomolecule structure. In a variety of examples we show how different molecular interactions determine the mechanical stability of polysaccharides, proteins, DNA and other biomolecules.

Under the influence of a stretching force of around 750 pN the polysaccharide dextran undergoes a transition into a stretched conformation during which bond angles flip into a new conformation. Molecular dynamics simulations corroborated this conformational change.

Proteins fold into compact domains. Especially for structural proteins the mechanical stability of these domains is of importance. We show that the immunoglobulin and fibronectin III domains of the muscle protein titin exhibit exceptionally high unfolding forces (150-250 pN) when stretched at speeds of normal muscle operation (Fig. 1).
3d Confocal Microscopy and Image Analysis for Measurement of Genetic Instability
Authors: Ortiz de Solorzano C , Chin K , Knowles D , Jones A , Garcia E , Gray JW , Lockett SJ
Published: July 2020

Solid tumors frequently contain cells that are heterogeneous in the copy number of DNA loci. This fact implies the existence of genetic instability, which may be associated with disease aggressiveness. Accurate measurement of this phenomenon requires analysis of intact nuclei within their natural tissue context. We perform these measurements by preparing >30μm thick tissue sections, labeling them with fluorescent labels for total DNA and for specific DNA loci using fluorescence in situ hybridization (FISH) which retain the transparency of the tissue and acquiring 3D images of the tissue using confocal microscopy (figure 1). In this study, we combined automated 3D image analysis (IA) algorithms for segmenting individual nuclei based on the total DNA stain1 and for segmenting the punctuate FISH signals of DNA loci. This enables us to efficiently enumerate the copy number of specific DNA loci in individual cells and as a function of the cell's location in the tissue.
Monitoring of Cell Surface Movement at the Erythrocyte’s Edge By Scanning Phase Contrast.Microscopy
Authors: Bitler Arkady , Barbul Alexander , Korenstein Rafi
Published: July 2020

We explored the ability of a novel method of nano-scale movement detection to analyze the effects of chemical crosslinking and thiol reduction on the fluctuations of the human erythrocyte membrane. The method employs line images of light intensity distribution across the red blood cell, obtained by laser scanning phase contrast microscope. The essential feature of these images is a noiseless region corresponding to edge scene. This enables- one to carry out very accurate fitting procedure between the theoretical intensity distribution and the experimental one. One of the free parameters of fitting is the edge position. Measuring line scan intensity distributions with a certain sampling rate and defining the edge positions in consequent moments of time, yields the time-dependent displacement of cell's edge with up to nanometer scale precision.

The significant feature of the suggested procedure is that it implicitly takes into account the halo effect.
Early “Events” in Hiv Infection and Hiv-Associated Neoplasia
Authors: Brian Herndier
Published: July 2020

The course of HIV infection necessarily involves the initial establishment of the virus in a receptive host environment. HIV disease involves active intracellular replication of the virus and spread of the virus from the initial compartment of the infection. Finally, from the virus’ perspective _ virions must enter a compartment suitable for eventual host-host transmission. In the vernacular of HIV epidemiology this implies some sort of “exchange of bodily fluids”.

Recently, our research focus has been on tracking HIV during the early phases of infection. The goal is to piece together a full chronology of the disease at the tissue level. The emphasis will be on changes in lymphoid architecture that might ultimately be important in the pathogenesis of AIDS malignancies and offer insight into the dysfunctional aspects of immunity in the setting of retroviral infection. Because HIV disease often ultimately features a severe immunodeficiency, other pathogens by necessity play a role in shaping the “visible” immune response.
Aids/Opportunistic Infections
Authors: Guccion J
Published: July 2020

Use of TEM in the diagnosis of opportunistic infections in AIDS patients is indicated in three situations. First, when an opportunistic infection is suspected, but an infectious agent is not visible in tissue sections studied by light microscopy (LM). This situation is illustrated by the following case.(1) A 33-year-old man with AIDS presented with fever, nightsweats, generalized lymphadenopathy, and a 30 lb. weight loss. An inguinal lymph node biopsy was performed. There was no consensus as to the diagnosis among the pathologists who studied H&E sections of the biopsy. An obscure infectious disease, a B-cell lymphoma with amyloid stroma, and possibly Hodgkin's disease were the major considerations. TEM study revealed unexpected clusters of small, pleomorphic, coccobacillary bacteria in vascular walls typical of cat scratch disease. These bacteria were later further identified by polymerase chain reaction as belonging to the genus Rochalimaea(now Bartonella), an important cause of cat scratch disease.
Aids-Associated Viral Infections
Authors: Miller Sara E
Published: July 2020

Infection with human immunodeficiency virus (HIV) eventually causes a profound decrease in the body's ability to eradicate or control infections with microorganisms, including viruses. Some infections in AIDS patients are due to common organisms which are of little significance in immunocompetent individuals. Other organisms can be harbored continuously, occasionally causing disease, but normally being suppressed after a heightened immune defense; in AIDS patients, these infections can be life-threatening. Further, practices that predispose to HIV infection also permit entry of other organisms, such as hepatitis and herpesviruses. Electron microscopy is beneficial as an adjunct to other modalities for viral detection. Methods for identifying viruses, both in fluids by negative staining and in tissues by thin sectioning, have been published. Some viral pathogens, including HIV itself, are best documented by other means.

HIV has been demonstrated by EM in infected individuals, but because it destroys and makes scarce the cells for which it has an affinity, it is difficult to find them.
Permeability Oxygen Through Endothelium By Time-Resolved Fluorescence Microscopy : Influence.of Plasma Viscosity on Tissue Hypoxia.
Authors: Dominique Dumas , Véronique Latger , Viriot Marie L , Stoltz Jean F
Published: July 2020

Vascular endothelial cells (EC) forms a continuous monolayer, a selective barrier that coats the inner surface of the vascular system, representing the primary anatomical site between blood and tissue. Under closed conditions (shear stress, hypoxia-reoxygenation..), EC are activated and the level of adhesion molecules (ICAM-1) expression increased. A technique of fluorescence quenching by oxygen in cell membrane was used to study molecular transport processes on a larger scale1-4. Collisional fluorescence quenching is described by the Stern-Volmer equation : τ0 / τ = 1 + kd.PO2 where τ0 and τ are respectively the excited state lifetimes in the absence and in the presence of oxygen, Kd the quenching dynamic constant and PO2 the oxygen partial pressure. The main purpose of the present study was to investigate the relation a) between the plasma viscosity and PO2 across human EC monolayer by Time-Resolved Fluorescence Microscopy (TRFM5-6 ) between the PO2 calculated and the level of ICAM-1 expression by flow cytometry (FCM)7 and Conventional Optical Scanning Microscopy (COSM8).
In Vivo Characterization of Cardiac Tissue Defects Associated With Pdgfrα Gene Deletion In the Patch Mouse
Authors: Thielen T , Terracio L , Borg TK , Price RL
Published: July 2020

The active form of platelet derived growth factor (PDGF) is a polypeptide dimer of which there are three isoforms: PDGF-AA, PDGF-BB, and PDGF-AB. Furthermore, two types of PDGF receptors also exist: PDGFRα and PDGFRβ. The β-receptor binds only the PDGF-B chain with high affinity, while the αreceptor can bind either PDGF-A or -B chains. Both PDGF ligands and their receptors have been linked to such cellular responses as mitogenisis, cell migration, chemotaxis, adhesion and differentiation in a variety of cell types.

Several studies have reported fetal cardiac abnormalities associated with disruption of PDGF and its receptors. In spontaneous mutants lacking the PDGF α-receptor (Patchmice), fetal hearts are generally dilated and exhibit valve and outflow tract defects. Similar defects are observed in normal embryos injected with antibodies to block the functioning of the PDGF-A chain. Disruption of PDGF-B function results in ventricular and atrial dilation, reduction of the ventricular wall, and hyper-trabeculation.
Biological Relevancy of Quantitative Analysis of Adherence Receptors on Polymorphonuclear.Neutrophil by Flow Cytometry and Optical Scanning Microscopy..
Authors: Dumas Dominique , Latger Véronique , Stoltz Jean F
Published: July 2020

Leukocytes adherence receptors are the basis of cells interactions1. Flow cytometry (FCM) makes it possible to assess a characterization of the activation level of a given cellular population by receptor quantifying2. That technique, however does not integrate at the same time, all others factors of leucocyte adhesive phenotype regulation, as spatial distribution and molecular conformation. Our study consisted in exploring the main adhesion receptors (CD62L, CDllb/CD18) on the surface of Polymorphonuclear Neutrophils (PMN) that were prepared under identical conditions and simultaneously analyzed by FCM and Conventional Optical Scanning Microscopy/Deconvolution3 (COSM). Methods: PMN cells from whole blood were obtained using Polymorphprep™. Isolated PMN (105 cells /ml) were incubated with Tumor Necrosis Factor (TNFα, 100 UI/ml) at 37°C for one hour. Quantification of cell surface molecules was assessed according to the quantitative indirect immunofluorescence3 method using QIFIKIT® (Dako, France).
The Structure of Maize Streak Virus: Are Two Heads Better Than One?
Authors: Zhang W , Olson N H , Chipman P R , McKenna R , Baker T S
Published: July 2020

The geminiviruses are a group of plant DNA viruses characterized by having one or two copies of a single-stranded, circular DNA genome and frequent occurrence of geminate, quasi-isometric particles as observed by transmission electron microscopy. The Geminiviridaefamily is classified into three genera (Mastrevirus, Curtovirus and Begomovirus)based on the number of DNA components, the plant hosts, and the transmission vectors. Maize streak virus (MSV), the type member of the Mastrivirusgenus, is transmitted by the leafhopper and infects Gramineaespecies. Wheat dwarf virus, Digitaria streak virus, and Chloris striate mosaic virus (CSMV) are examples of other members of the Mastrivirusgenus. The Nigerian strain of MSV (MSV-N) contains one copy of a singlestranded, circular DNA molecule (2687 nucleotides). Two protein products, VI and V2, are coded by the MSV-N genome.2 V2 (244 amino acids) is the only coat protein found in virions.
Ultrastructural Diagnosis of Whipple& Disease at Autopsy
Authors: Siew S , Newton B
Published: July 2020

Whipple's disease is a rare systemic disorder with the intestinal tract being the major site of predilection. The characteristic histologic features are the infiltration of large numbers of macrophages, which have a clear or vacuolated cytoplasm. In his original documentation of “a hitherto underscribed disease”, Whipple interpreted the vacuo lation to be due to deposits of fat and fatty acids and gave the name “interstinal lipodystrophy” to the pathology. In this report, Whipple did describe the presence of rod shaped organisms in Levaditi (reduced silver) stained preparations, but, it was not until a half century later, that the definitive presence of intracellular organisms was demonstrated by means of transmission electron microscopy. Further investigations showed that the organism is a Gram positive actinomycete, which has been designated as Tropheryma whippelii.

We report the case of a 72 year old woman, who was admitted with the chief complaints of abdominal cramps and diarrhea.
Centrosome and Centriole Abnormalities During Cancer In the Transgenic Adenocarcinoma Mouse Prostate (Tramp) Model
Authors: Schatten H , Wiedemeier A , Taylor M , Lubahn D , Greenberg N M , Besch-Williford C , Rosenfeld C , Day K , Ripple M
Published: July 2020

Centrosome abnormalities and abnormal mitotic formations have been shown in human breast cancer tissue and in LNCaP and DU145 prostate cancer cells in culture. Here we report on abnormal centrosome and centriole organization in the transgenic adenocarcinoma mouse prostate (TRAMP) model and on the implications for abnormal cell divisions, genomic instability, and apoptosis. Centrosomes are microtubule organizing centers which assemble bipolar spindles in normal cells. In cancer, centrosomes cause the formation of mono-, tri-, and multipolar mitoses which will separate chromosomes unequally during cell division.

We investigated the occurence of abnormal mitoses in TRAMP tissue in different stages of cancer progression using transmission electron microscopy (TEM). Advanced stages of prostate cancer are characterized by multiple abnormal mitoses ranging from tripolar to unusual multipolar formations. In addition, apoptotic bodies are abundant which may indicate that multipolar mitoses will lead to cell death or apoptosis.
Cardiac Rhabdomyoma: Ultrastructural Features and Review of Literature
Authors: Zhu Shen-Hua , Barrish James P , John Hicks M
Published: July 2020

Primary cardiac tumors of any type in infancy and childhood are rare, with a prevalence of 0.03%reported in autopsy surveys carried out at children's hospitals. Within the pediatric population,the most common primary tumor involving the heart is rhabdomyoma. These tumors may bedivided into 3 groups: 1) arising in tuberous sclerosis; 2) arising sporadically; or 3) arising in congenital heart disease. Many cardiac rhabdomyomas are asymptomatic and regress early in life without clinically untoward effects. Only a small proportion of these tumors come to the attention of the pediatrician and pathologist when there is obstruction of blood flow, a murmur related to subaortic stenosis, or arrhythmias. Rhabdomyomas may be detected prenatally during ultrasound examination, with one case being diagnosed at 22 weeks gestation. Although it is possible to subdivide the tumors into 3 groups, the association of this entity with tuberous sclerosis is very strong with at least 60 to 85% of children with rhabdomyomas having tuberous sclerosis.
Microcysys In the Dorsal Cochlear Nucleus of the Acoustically Deprived Gerbil
Authors: Yu SM , Yang YJ
Published: July 2020

It has been shown that cavities or vacuoles are most evident in the posteroventral and antero ventral cochlear nucleus (PVCN & AVCN) of the gerbil. These vacuoles are reported as a neurodegenerative disorder or spongioform degeneration or a dynamic process related to the degree of auditory stimulation or microenvironmental homeostasis in the cochlear nucleus of the gerbil. Little information is avalable on the origin and formation of microcysts in the dorsal cochlear nucleus (DCN) of the gerbil. The aim of this study, was to investigate the possible formation of microcysts in the gerbil DCN after a surgical ligation of the external auditory canal by transmission electron microscopy.

The mongolian gerbil, Meriones unguiculatus,had been acoustically deprived on the right side by a surgical ligation of the external auditory canal at six months old.
Ultrastructural Characterization of Hearts from Transgenic Mice Expressing the Epstein-Barr Virus Nuclear Antigen-Leader Protein
Authors: Megill JR , Monticello T M , French MH , Smith PL , Trippodo NC , Durham SK
Published: July 2020

Mice expressing the Epstein-Barr virus nuclear antigen-leader protein (EBNA-LP) develop congestive heart failure and atrial thrombosis as early as four months of age. Epstein-Barr virus (EBV) is a common human herpes virus with a complex life cycle involving chronic replication in epithelial tissues. The EBNA-LP, one of six EBV nuclear antigens, has an unknown biochemical function but is associated with EBV-induced growth transformation. This transgenic line was originally produced to investigate oncogenic consequences, but unexpectedly resulted in mice with progressive heart failure. Since the progression and development of the heart failure is highly predictable, this transgenic line may serve as a valuable model for studying the pathophysiologic changes associated with human dilated cardiomyopathy. We describe here the ultrastructural characteristics of the myocardium from EBNA-LP transgenic mice in order to better delineate the pathogenesis of the associated heart disease.
Sem And Tem Observations of Rat Trachea and Lung After Inhalation Injury and Cutaneous Burn In a Rat Model
Authors: Langlinais PC , Mozingo DW , Dubick MA , Carden SC , Goodwin CW
Published: July 2020

Inhalation injury is present in 32-38% of patients with severe burns and is associated with an increase of 20-84% above the mortality expected based on age and burn size alone. Most previous studies of smoke inhalation injury have utilized large animals such as the sheep and we have previously reported a TEM and SEM study of lung injury in the sheep. The present observations are part of a study to develop a small animal combined model of smoke inhalation and surface burn.

Adult, male Sprague-Dawley rats were used. Animals were anesthetized and randomly assigned to one of four groups. Groups 1 and 2 received a 20% total body area surface (TBSA) full thickness scald burns while groups 3 and 4 were sham treated. Five hours after burn injury, rats were placed in a nose only exposure device and half of each group was exposured to either room air alone or room temperature tree bark smoke for 16.25 minutes.
The Vascular Structure of the Kidney of Mallards and Doves
Authors: Ditrich H
Published: July 2020

The architecture of the kidney of birds (and also reptiles) is, unlike in mammalians, mainly determined by the organization of the blood vascular system. Besides arterial supply and venous drainage, the renal portal system forms a main structural component. While the latter was often regarded as a “primitive” feature in the literature, morphological and physiological data reveal its great functional importance.

Microvascular corrosion casts studied in the scanning electron microscope permit the visualization of minute vessels, retaining their 3D-arrangement. Additionally, when compared with graphical reconstructions of serial sections, this method avoids several inherent artifacts like fixation and dehydration shrinkage as well as the compression of the object by the sectioning blade. Most of the studies on avian kidneys with this technique used the domestic chicken as a model. In order to provide additional material for comparative and functional studies, data on the intrarenal vascular structure of other species are required.
Blood Supply and Function of the Tegmentum Vasculosum (Stria Vascularis) of the Duckling Cochlea
Authors: Hossler FE
Published: July 2020

On the basis of ultrastructural, biochemical, cytochemical, and electro-physiological evidence, the stria vascularis, a highly vascular epithelium of the mammalian cochlea, has been implicated as one site of endolymph synthesis. It would seem natural to suspect that its highly developed counterpart in the avian cochlea, the tegmentum vasculosum (TV), might have a similar function. This study utilizes routine light and electron microscopy, biochemistry, cytochemistry, and vascular corrosion casting to describe the structure and blood supply to the (TV) in the duckling and provide evidence for its involvement in endolymph synthesis.

Four to 16 week-old ducklings (Anas platyrhyncos)were anesthetized and anticoagulated and perfused via the common carotid arteries with buffered, 2% glutaraldehyde for routine light microscopy or transmission or scanning electron microscopy, or with Mercox resin for vascular corrosion casting. Casts were macerated in 5% KOH and warm water to remove tissue, cleaned in formic acid and water, dried by lyophilization, mounted on stubs and viewed at 10-20 kv by SEM.
Comparative 3d-Morphometry of the Microvascular Unit In Tumors and Normal Tissues: Quantitative Studies on Corrosion Casts
Authors: Konerding M A , Malkusch W , Presta M , Gaumann A
Published: July 2020

The significance of angiogenesis and the vascular architecture have been stressed in numerous studies of the structure and biological properties of the tumor vasculature and blood flow (Folkman and D'Amore, 1996; Jain, 1988). The important question as to whether the vascular architecture of an individual tumor is tumor type specific has been controversial. This is due, at least in part, to the methodologies used. Most reports confine themselves to qualitative observations and comparisons of gross vascular patterns in host and tumor, or to blood vessel density, length and diameter measurements, which in turn vary with the staining and counting techniques.

We studied the microvascular architecture of four different tumor cell lines (CaX, CaNT, SaS, HEC- 1B) in order to determine whether there is a characteristic vascular pattern for different tumor types and whether it differs significantly from two normal tissues, muscle and gut.
Microstructural Changes of the Compact Bone in the Mandible of Aging Rats Using Plastic Casting Method
Authors: Okada S , Iwaku F , Yoshida S , Ohta Y
Published: July 2020

A method for producing plastic castings of the internal structure of hard tissue has been described by Marten et al. (1978). Although they examined dentine, this technique could also be used to examine other hard tissue such as bone (Weber, 1983). The aim of our study is to expose the true threedimensional relationships between the internal structures of the compact bone and these microstructural changes of the mandible in the aging rats.

Nine Wistar male rats were used in this study and divided into 3 groups as follows: 3 weeks-old rats as young stage, 12 weeks-old as adult stage and 80 weeks-old as old stage. All rats were perfused with 2.5% glutaraldehyde via the ascending aorta. In one rat for each group, the mandible was decalcified by 10% EDTA and embedded in paraffin. 10 μm sections were stained with Schmorl’s staining method.
Self-Limiting Aggregation By Controlled Ligand-Receptor Stoicfflometry and Its Use For a Novel Drug Delivery System
Authors: Kisak E , Kennedy M , Zasadzinski J A
Published: July 2020

Lipid vesicles are used as drug delivery vehicles for the slow sustained release of a drug compound to a specific site in the body. This translates to more efficient medication with limited side effects. Although unilamellar drug delivery vesicles have progressed greatly, they are still limited in there applications. Our group has designed a second generation drug release system, the “vesosome“ which incorporates an aggregate of lipid vesicles encapsulated in a second lipid membrane. The two separate membranes can be specialized to allow for increased drug encapsulation and better control over drug release rate, which leads to a more general drug delivery system.

Lipid vesicle aggregates were formed by using a ligand-receptor system (biotinated lipids protruding from the vesicle surface crosslinked with streptavidin). The streptavidin/biotin system is one of the strongest in nature, providing specific binding.
Ultrastructural immunocytochemical Synapse Changes In A Rat Model Of Parkinson’s Disease
Authors: Meshul CK , Allen C , Kay T S
Published: July 2020

Parkinson's disease is a progressive disorder that is characterized by degeneration of the dopamine containing neurons located within the midbrain (substantia nigra). There is also substantial loss of dopamine within nerve terminals located within the striatum which originate from those dopamine neurons. Current therapy involves administration of the precursor to dopamine, namely l-dopa. This chemical is taken up into the brain and then converted to dopamine. Although replacement of dopamine is effective over the first few years, other movement disorders are associated with longterm l-dopa therapy. The l-dopa induced dyskinesias limit the usefulness of this type of therapy.

Although loss of dopamine is the major neurochemical deficit in Parkinson's disease, other neurotransmitters within the striatum may also be altered. There is a major axonal projection from the cortex to the striatum. The corticostriatal pathway uses the excitatory neurotransmitter, glutamate, and dopamine is known to modulate the activity of glutamatergic synapses
Alterations In Glutamate Synapses In The Nucleus Accumbens Following Sensitization To Cocaine
Authors: Kozell L B , Meshul C K
Published: July 2020

Repeated intermittent administration of cocaine to rodents has been shown to result in behavioral sensitization, or enhanced locomotor activity. Because there are similarities between effects of intermittent cocaine use in humans and sensitization in rats, understanding the neural mechanism underlying sensitization may provide insights into mechanisms of psychopathologies in humans.

There is abundant evidence that the mesolimbic dopaminergic pathways, originating in the ventral tegmental area (VTA) and terminating in both nucleus accumbens (NAc) and prefrontal cortex (PFC), are critically important for development of sensitization to the motor-stimulating effects of cocaine. Evidence also suggests that changes in glutamate (Glu) transmission in the mesolimbic DA pathways are associated with sensitization to cocaine. In recent collaborative studies, we have shown that cocaine administration transiently decreases the density of nerve terminal glutamate immunolabeling within the NAc shell following withdrawal from continuous or intermittent cocaine administration. Locomotor activity was not assessed in either of these studies.
Interactions Between Mouse Lymphocytes And Borrelia Burgdorferi, The Infectious Agent Of Lyme Disease
Authors: Dorward D
Published: July 2020

Lyme disease is a tick borne, multi-system disorder caused by low density systemic infections with the spirochete Borrelia burgdorferi. Without antimicrobial treatment, mammalian infections with these bacteria are persistent and chronic. Recent studies showed that B. burgdorferi can target, invade, and lyse both cultured and primary human B and T cells. Direct interactions between the spirochetes and lymphocytes also leads to adherence of B and T cell antigens on the surface of significant proportions of the bacteria. Adherent lymphocytic antigens inhibit binding of antibodies to prominent B. burgdorferi proteins, and interfere with classic complement-mediated killing, suggesting a possible role for spirochete-lymphocyte interactions in immune evasion.

In order to develop an experimental animal model for assessing spirochete-lymphocyte interactions, B. burgdorferi and primary mouse lymphocytes were co-incubated and examined by electron microscopy. Mononuclear cells were separated from fresh mouse blood by Ficoll gradient centrifugation (ICN, Biomedicals, Aurora, Ohio).
Correlative Sem And Tem Of Plant Spermatozoids: Equisetum arvensel.
Authors: Schmitt S J , Renzaglia K S
Published: July 2020

The phylogenetic position of mosses, ferns and fern allies is enhanced by detailed knowlegde of gametophyte reproductive structures, in particular the spermatozoid (freeswimming sperm). The number and placement of flagella in conjunction with their ultrastructure and that of the multilayered structure (MLS) has been used to establish related groups. Herein, ultrastructural details of pre-release spermatozoids obtained via TEM are compared with the morphological appearance of fully mature (free-swimming) spermatozoids obtained via SEM. This represents the first intensive study of any plant sperm cell that correlates 3-dimensional SEM images with TEM ultrastructural features.

Equisetum arvense L. is a fern ally commonly refered to as ‘horsetail’. The mature spermatozoid (Fig. 1) is twisted into approximately 3.5 revolutions, or gyres. A complex locomotory apparatus (Fig. 2), involving approximately 55 flagella, dominates the anterior region of the cell. The basal bodies subtending these flagella feature a well-defined stellate pattern and occur in 4 irregular rows staggered over approximately 1.75 gyres.
Speedy Plankton: Myelinated Axons In Calanoid Copepods (Crustacea)
Authors: Davis April D , Weatherby Tina M , Lenz Petra H
Published: July 2020

Copepods are an abundant and diverse group of crustaceans. One order of free living copepods, the calanoids, are usually < 3 mm in length, planktonic, and possess very rapid escape responses. These animals dominate planktonic communities and their escape reactions contribute to their success. Although all calanoids respond to hydrodynamic disturbances with an escape jump, minimum reaction times vary among species, ranging from 1.5 to 6 msec. To help us understand the physiological basis underlying the phenomenally short reaction times we initiated a comparative study of the internal structures by using transmission electron microscopy (TEM).

Traditional chemical fixation was adequate for several species of copepods, and sensory structures were clearly shown. In these calanoids, minimum reaction times ranged from 3 msec to 6 msec and can be partially explained by giant axons, the most common way invertebrates increase the rate of conduction of nerve impulses. However, the internal structures of two copepod species, Euchaeta  rimana and Undinula vulgaris, were distorted (Fig. 1).
Quantitative 3d Analysis Of Intra-Nuclear Organization In The Tissue Context
Authors: Lockett SJ , Knowles DW , Pinkel D , Ortiz de Solórzano C
Published: July 2020

Confocal microscopy is revealing associations between the internal organization of the nucleus and tissue architecture and function. Such associations may exist which are too subtle or complex for visual observation or quantitative analysis may be required, for example as input data to mathematical modeling of cellular processes. In these circumstances, it is necessary to perform the analysis using computer algorithms. We have developed 3D image analysis (IA) algorithms for segmenting nuclei from within intact tissue specimens, measuring the structure of the nuclei and for segmenting specifically- labeled punctate entities within nuclei. In this study we developed algorithms for measuring the spatial organization of the two copies of a specific DNA locus (labeled using fluorescence in situ hybridization (FISH) ) inside diploid nuclei and with respect to the nuclear organization of the tissue.

For each segmented nucleus, its center of mass (CoM) was determined, which informed us about its position in the tissue.
The Role Of P115 Rhogef In The Rho Mediated Effects Of Lpa On The Actin Cytoskeleton.
Authors: Schwartz CL , Wells C , Jiang X , Arnott HJ , Sternweis PC , Wilk-Blaszczak M A
Published: July 2020

In the fibroblast cell line, 3T3, lysophosphatidic acid (LPA) induces stress fiber formation. Stress fibers participate in physiological functions such as cell motility. LPA acts through a receptor coupled to a PTX-insensitive G-protein, G13. It was shown that a constitutively activated mutant of α13 (Q226L) induces stress fiber formation in Swiss3T3 cells through a second messenger cascade that involves a monomeric G-protein, Rho. The recently discovered guanine nucleotide exchange factor, p115 RhoGEF (p115) forms a link between a n and Rho A. In the presence of α13, p115 activates Rho. The N-terminus of p115 contains a regulator of G-protein signaling (RGS) box. RGS proteins act as negative regulators of G-protein dependent signaling by increasing GTPase activity and “locking” the G-protein in an inactive state. We have tested a role of p115 in the pathway coupling the LPA receptor to stress fiber formation by Rho in NTH-3T3 cells.
25 Years In A Stereology Laboratory: A Point By Point History
Authors: Basgen John M
Published: July 2020

In many experimental situations there is some three dimensional particle that is of interest to the microscopist. To obtain morphometric information about the particle, the microscopist must section some reference space that contains the particle or component of interest, but after sectioning, the reference space and component are reduced to a 2-D sample. This sample no longer contains 3-D structures but only 2-D profiles of the reference space and component.

Stereology can be used to obtain 3-D information from 2-D samples. These samples can be micrographs or digital images, or they can even be the cut surface of the 3-D reference space. The four most basic parameters measured using stereological techniques are volume (3-D), area (2-D), length (1-D), and number (0-D). When measuring these parameters it is often advisable to first measure the density of the component within the reference space, next measure the volume of the reference space, and finally multiply the density by the reference volume to obtain the actual structural parameter.
High Speed Two Photon Excitation Microscopy in Live Cell Imaging using Image Correlation Spectroscopy (ICS)
Authors: Wiseman P W , Bouwer J C , Peltier S , Ellisman M H
Published: July 2020

For live-cell imaging, two-photon excitation microscopy (TPEM) is proving to be a significant technological advancement. The unique features offered by TPEM are the ability to image thick sections, excellent optical sectioning capabilities, low damage to living cells, and less out of focus fluorescence and out of focus photobleaching. of these features, the most useful for the biological microscopist, is optical sectioning. Optical sectioning is an intrinsic property of the two-photon process, whereby, two infrared (IR) photons are absorbed quickly to excite a single UV/blue transition. The probability for exciting a two photon transition is proportional to the instantaneous excitation intensity squared. Therefore, for a focused laser beam, only light at the focal point of the excitation beam excites a fluorescent transition. Thus, the need for confocal apertures and time consuming deconvolution algorithms are, for the most part, eliminated.

We have continued to develop and enhance our ability to perform high-speed, two-photon excitation fluorescence microscopy. in 1998, we successfully deployed a prototype, video-rate twophoton laser scanning system (30 frames/sec or faster at reduced scan width) developed with support from Nikon Corporation. That system was built upon a Nikon RCM 8000 confocal microscope.
Study of the Growth Curve of Vero Cell with Statistical Counting Method Based on the SEM Images
Authors: Shao Manjun , Jiang Lei
Published: July 2020

For industrial-scale culture of anchorage-dependent mammalian cell, such as vero cell, growth on the microcarriers offers certain advantages. As previous research reported, growth process can be illustrated by a growth curve - cell density versus time, which shows the change of cell density through whole growth process including: first adhesion stage; spreading and division stage; and last confluent stage. The growth curves are controlled by the culture condition (culture medium, temperature, pH value, inoculation concentration, etc.) and space condition (kind of microcarrier, microcarrier size, and surface property, etc.). Therefore, the cell growth curve is not only the important basis for studying growth kinetics, but also for designing biological reactors.

Traditionally, total cell concentration for the microcarrier cultures were determined using the crystal violet method, and the cell nuclei were counted with a hemacytometer. in this method, the cells attached on each microcarrier were trypsinized and dyed by crystal violet dye, so that the number of nuclei counted was statistically representative. in this work, the cells and microcarriers were examined by using an environmental SEM - KYKY1500. As a result, the moist cells and microcarriers in SEM images show their natural characteristics, because this environmental SEM permits biological sample to be observed directly in an environmental specimen chamber without normal preparation for biological sample. As for the normal preparation, it needs a long duration, e.g. 2 days normally, to do fixing, desiccating and gold-coating. in contrast, the operation for observation of the biological samples in this SEM is much simpler. in view of this, a statistical counting method based on the SEM images for determining cell concentration during microcarrier culture was developed.
The Glomerulus-Revisited
Authors: Siew S
Published: July 2020

In the embryologic development of the nephron, the glomerular tuft (renal corpuscle) and the excretory tubules develop from two distinct embryologic origins or anlagen. Nephrotomes constitute the earliest formation of the excretory tubules and appear first as segmentally arranged cell clusters, which grow and become canalized, forming nephric tubules. Formation of the glomerulus begins when the dorsal aorta gives rise to small branches, consisting of tufts of capillaries. Bowman’s capsule forms as a blind terminal expansion at the proximal end of the nephron. in the final stage of development, Bowman’s capsule becomes deeply indented by the glomerular tuft. Which leads to a differentiation of it into 2 layers: 1) an inner, visceral layer and 2) an outer, or external, parietal layer. The two layers are separated by a crescent shaped space, known as Bowman’s or the urinary space.

The inner, visceral layer encloses, and, comes into close apposition with the glomerular capillaries. At the vascular pole of the glomerulus, the visceral layer is reflected off these vessels and becomes continuous with the external, or parietal layer of Bowman’s capsule, which is lined by a very flat polygonal epithelium. Traditionally, it has been depicted that there is a direct opening of the urinary space into the tubule and that the flat cells of the parietal layer of Bowman’s capsule become continuous with the cuboidal epithelium in the neck of the proximal convoluted tubule.

This investigation shows that the junction of the renal corpuscle and the proximal convoluted tubule is a more complicated one. Scanning electron microscopic studies of microdissected nephrons enabled the visualization of the glomerulus, with the two vessels at the vascular pole and the proximal convoluted tubule at the urinary pole. An unexpected finding, was the presence of a gourd-like out-pouching at the urinary pole. (Fig. 1, “A”) Higher magnification showed a saucer shaped depression (50 μm × 25 μm) on its free border. (Fig. 2). This was a consistent finding in the material that was studied.
Structure and Function of the Primary Zygote Wall of Chlamydomonas Monoica
Authors: Salanga M , Van Winkle-Swift K
Published: July 2020

The Chlamydomonas zygospore is formed after gametic cell fusion by a progression of ultrastructural and physiological changes, including the synthesis of a protective and elaborately sculptured secondary wall (Figure 1). Assembly of the secondary wall occurs within the confines of a transient primary zygote wall, synthesized immediately after gamete fusion, and assembled beneath the residual gamete walls. Upon completion of secondary wall assembly, the primary zygote wall is shed from the maturing zygospore (Figure 2). This primary zygote wall, composed primarily of a β-1,3 glucan (callose), may be important in the regulation of secondary zygospore wall formation.

A temperature-sensitive mutant strain of C. monoica, zym30, has been isolated that shows marked abnormalities in both primary and secondary wall ultrastructure. The degree of wall formation in the mutant strain is inversely proportional to temperature. At 25° C most zygotes fail to synthesize either the primary or the secondary wall. At 15° C both primary and secondary wall assembly occurs to some extent and as many as 50% of the zygotes develop a “pseudo-wildtype” phenotype (Figure 3). Although release of the primary wall rarely occurs in the zym30 strain (even at permissive temperatures), those walls that are released appear thinner than the walls shed from wildtype zygospores, as viewed by phase contrast microscopy. Fluorescence images incorporating aniline blue, a fluorochrome specific for β-1,3 glucans, show decreased fluorescence suggesting reduced callose synthesis (Figure 4).

Several permanent, ultrastructurally distinct layers comprise the secondary wall, the assembly of which may be controlled, in part, by the primary wall. One or more of these secondary wall layers contain the highly durable, autofluorescent biopolymer, sporopollenin. Autofluorescence of zym30 zygospores produced at the permissive temperature is enhanced relative to wildtype zygospores. This correlates with ultrastructural changes in the primary wall and overproduction and/or misassembly of the surface secondary wall layer (Figure 5). Work is in progress to determine whether the environmental resistance, and/or viability of the zym30 “pseudowildtype” zygospores are reduced relative to wildtype zygospores.
Increasing Access to Tomographic Resources: Web-based Telemicroscopy and Database
Authors: Martone M E , Peltier S , Lamont S , Gupta A , Ludaescher B , Khetani S , Molina T , Ellisman M H
Published: July 2020

The application of electron tomography to cell biology has led to important insights into the 3D fine structure of subcellular processes and organelles. Tomography has been particularly useful for studying relatively large, multi-component structures such as the Golgi apparatus, mitochondria and synaptic complexes. When combined with very powerful high voltage electron microscopes, tomography has also provided high resolution quantitative views of extended structures such as neuronal dendrites in very thick sections (4 μm) at electron microscopic resolution. The utility of tomography is twofold: first, it provides 3D examination of subcellular structure without the need for serial section analysis; second, because the computed slices through the tomographic volumes can be much thinner than is possible to produce by physical sectioning, it reveals structural detail in the range of 5-30 nm that tends to be obscured in conventional thin sections. Tomographic analysis has forced re-assessment of long-standing views of organelles such as mitochondria and the Golgi apparatus and as the technique advances, additional insights are likely forthcoming.

Electron tomography is an expensive technique, both in terms of the instruments used and the computational resources required. The three major high voltage electron microscope resources in the United States, San Diego, Boulder and Albany, all are actively engaged in tomographic research and offer this important technology to the scientific community at large.