Digital light microscopy as a tool in toxicological evaluation of fish erythrocyte morphological abnormalities

2020 ◽  
Vol 83 (4) ◽  
pp. 362-369 ◽  
Author(s):  
Eldores Sula ◽  
Valbona Aliko ◽  
Maria Pagano ◽  
Caterina Faggio
Keyword(s):  
Light Microscopy ◽  
Toxicological Evaluation ◽  
Morphological Abnormalities ◽  
Fish Erythrocyte

Congenital Dyserythropoietic Anemias: Molecular Diagnosis and Diagnostic Approach in a Cohort of Italian Patients

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2142-2142
Author(s):  
Veronica Agrigento ◽  
Sclafani Serena ◽  
Paolo Rigano ◽  
Rita Barone ◽  
Giuseppina Calvaruso ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Bone Marrow ◽  
Iron Overload ◽  
Light Microscopy ◽  
Peripheral Blood ◽  
Ineffective Erythropoiesis ◽  
Blood Samples ◽  
Morphological Abnormalities ◽  
Erythroid Hyperplasia ◽  
Sds Page

Abstract Introduction. Congenital dyserythropoietic anemias (CDAs) are hereditary rare erythropoietic disorder characterized by distinct morphological abnormalities of the bone marrow cells, ineffective erythropoiesis and systemic iron overload. This conditions is characterized by a maturation arrest during erythropoiesis with a reduced reticulocyte production in contrast with erythroid hyperplasia in bone marrow. Three types of CDA are known: types 1, 2 and 3.The identification of their causative genes provided evidence that these conditions have different molecular mechanisms that induce abnormal cell maturation and division. We describe the clinical and laboratory manifestations, the diagnosis procedure, the therapeutic approaches and the clinical phenotype in some cases of congenital dyserythropoietic anemia (CDAs) in Italian patients. The molecular analysis allow us to identify several genetic variants some of which have never been described previously. This report highlights the importance of recognizing CDAI even in countries where thalassemia is common. Among the different tools genetic study of CDA-related genes remains the main approach for pursuing the right diagnosis. Methods. Peripheral blood samples from 100 normal adults and 20 patients with different types of anemia were collected. Blood samples were analyzed by EMA binding test and by polyacrylamide gel electrophoresis in presence of sodium dodecyl sulphate ( SDS-PAGE). SDS PAGE was carried out in a 3.5-17% exponential gradient gel in Fairbanks buffer Genomic DNA was extracted from mononuclear cells of peripheral blood samples, by using a phenol-chloroform method. Polymerase chain reaction (PCR) products were sequenced directly using Big-Dye terminator 3.1 cycle sequencing kit and run on ABI PRISM 3130 DNA analyzer. The bone marrow of all patients was analysed with light or electron microscopy. The diagnosis of CDAs was based on the presence of mild to moderate anemia ineffective erythropoiesis and bone marrow erythroblasts morphological abnormalities. SEC23B, CDAN1, KLF1, BCL11A gene sequencing analysis was performed to highlight the presence of nucleotide variations and their relationship with the clinical presentation. Results and Discussion. We collected blood samples from 20 Italian patients with suspect of CDAs and 100 samples belonging to the healthy control subjects.None mutation in the genes analyzed was found in the samples controls.We identified SEC 23 B mutations in 4 out of 20 patients analysed. In two patients with suspect of CDAII we found two nucleotide variations; SDS PAGE in these patients identified the presence of abnormal band 3 and the EMA binding test resulted pathologic. Bone marrow (BM) light microscopy revealed more than 10% mature binucleated erythroblasts. In the patients suspected of CDA1, several gene variants were found in the CDAN1 gene, some described in the literature as mutations or polymorphisms, others of uncertain significance. In a patient diagnosed with CDA1 two novel mutations was found. EMA binding test resulted normal in all patients with CDA1. Light microscopy of the BM of CDA1 patients showed erythroid hyperplasia, presence of internuclear bridges between intermediate erythroblasts and characteristic pattern of spongy "swiss cheese" hetrochromatin in electron microscopy. None KLF1 and BCL11A mutations correlated with atypical CDAs was identified. In this last patients the EMA binding test resulted normal. CDAs are rare clinical hereditary disorders whose correct diagnosis is often delayed to adolescence or adulthood, when significant iron overload and end organ damage may have been occurred. Patients are often misdiagnosed with other congenital haemolytic anaemias. Inaccurate diagnosis can lead to inappropriate therapies, such as iron supplements, aggressive transfusion or splenectomy. However even if the gold standard for the CDAs diagnosis is the electronic microscopy, the identification of the mutated genes involved in the majority of CDAs patients made in recent years has improved the possibility of detecting these diseases. Disclosures No relevant conflicts of interest to declare.

The effects of stains and investigators on assessment of morphology of canine spermatozoa

1998 ◽  
Vol 34 (4) ◽  
pp. 348-352 ◽  
Author(s):  
MV Root Kustritz ◽  
PN Olson ◽  
SD Johnston ◽  
TK Root
Keyword(s):  
Light Microscopy ◽  
Phase Contrast ◽  
Eosin Y ◽  
Preparation Technique ◽  
Phase Contrast Microscopy ◽  
Morphological Abnormalities ◽  
Eosin B ◽  
Contrast Microscopy ◽  
Conventional Light Microscopy

Percentage and types of morphological abnormalities found in canine spermatozoa were evaluated by three investigators using three stains (Giemsa-Wright stain [Diff-Quik], eosin Y/nigrosin [Hancock], and eosin B/nigrosin [Society for Theriogenology morphology stain] with conventional light microscopy, compared to phase contrast microscopy on unstained samples. The percentage of spermatozoa with abnormal heads, midpieces, and tails varied by technique and by investigator. Average percentages of morphologically normal spermatozoa were significantly higher in samples stained with Diff-Quik and samples examined by phase contrast microscopy than in samples stained with Hancock or Society for Theriogenology morphology stains. No effect of investigator on the percentage of morphologically normal spermatozoa was assessed. Results suggest that staining or preparation technique may alter the morphology of canine spermatozoa artifactually.

Two- or three-way observations of the identical cell elements within the same sections by LM, TEM and/or SEM

1978 ◽  
Vol 36 (2) ◽  
pp. 82-83 ◽  
Author(s):  
Nakazo Watari ◽  
Yasuaki Hotta ◽  
Yoshio Mabuchi
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Fine Structure ◽  
Light Microscopy ◽  
Thin Sections ◽  
Transmission Electron ◽  
Identical Cell ◽  
Electron Microscopes ◽  
Scanning Electron

It is very useful if we can observe the identical cell elements within the same sections by light microscopy (LM), transmission electron microscopy (TEM) and/or scanning electron microscopy (SEM) sequentially, because, the cell fine structure can not be indicated by LM, while the color is; on the other hand, the cell fine structure can be very easily observed by EM, although its color properties may not. However, there is one problem in that LM requires thick sections of over 1 μm, while EM needs very thin sections of under 100 nm. Recently, we have developed a new method to observe the same cell elements within the same plastic sections using both light and transmission (conventional or high-voltage) electron microscopes.In this paper, we have developed two new observation methods for the identical cell elements within the same sections, both plastic-embedded and paraffin-embedded, using light microscopy, transmission electron microscopy and/or scanning electron microscopy (Fig. 1).

The Broad Applications of the Scanning Electron Microscope

1969 ◽  
Vol 27 ◽  
pp. 20-21
Author(s):  
C. T. Nightingale ◽  
S. E. Summers ◽  
T. P. Turnbull
Keyword(s):  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Light Microscopy ◽  
Surface Topography ◽  
Great Depth ◽  
Resolving Power ◽  
Depth Of Field ◽  
Pictorial Representations ◽  
Scanning Electron

The ease of operation of the scanning electron microscope has insured its wide application in medicine and industry. The micrographs are pictorial representations of surface topography obtained directly from the specimen. The need to replicate is eliminated. The great depth of field and the high resolving power provide far more information than light microscopy.

Tumor Diagnosis

1982 ◽  
Vol 40 ◽  
pp. 262-265
Author(s):  
Bruce Mackay
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Differential Diagnosis ◽  
Light Microscopy ◽  
Clinical Examination ◽  
Ultrastructural Study ◽  
Diagnostic Procedures ◽  
Specific Diagnosis ◽  
Transmission Electron ◽  
Microscopic Diagnosis

The broadest application of transmission electron microscopy (EM) in diagnostic medicine is the identification of tumors that cannot be classified by routine light microscopy. EM is useful in the evaluation of approximately 10% of human neoplasms, but the extent of its contribution varies considerably. It may provide a specific diagnosis that can not be reached by other means, but in contrast, the information obtained from ultrastructural study of some 10% of tumors does not significantly add to that available from light microscopy. Most cases fall somewhere between these two extremes: EM may correct a light microscopic diagnosis, or serve to narrow a differential diagnosis by excluding some of the possibilities considered by light microscopy. It is particularly important to correlate the EM findings with data from light microscopy, clinical examination, and other diagnostic procedures.

Effects of Hyperoxia on Lung Utrastructure

1970 ◽  
Vol 28 ◽  
pp. 26-27
Author(s):  
Gladys Harrison
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Light Microscopy ◽  
Oxygen Effects ◽  
Age Dependent ◽  
The Central Nervous System ◽  
The Subject ◽  
Space Age ◽  
Alveolar Septa ◽  
Extensive Damage

With the advent of the space age and the need to determine the requirements for a space cabin atmosphere, oxygen effects came into increased importance, even though these effects have been the subject of continuous research for many years. In fact, Priestly initiated oxygen research when in 1775 he published his results of isolating oxygen and described the effects of breathing it on himself and two mice, the only creatures to have had the “privilege” of breathing this “pure air”.Early studies had demonstrated the central nervous system effects at pressures above one atmosphere. Light microscopy revealed extensive damage to the lungs at one atmosphere. These changes which included perivascular and peribronchial edema, focal hemorrhage, rupture of the alveolar septa, and widespread edema, resulted in death of the animal in less than one week. The severity of the symptoms differed between species and was age dependent, with young animals being more resistant.

Immunocytochemistry. A Review of Methodology for Electron Microscopic Applicaiton

1974 ◽  
Vol 32 ◽  
pp. 328-329
Author(s):  
Joseph E. Mazurkiewicz
Keyword(s):  
Electron Microscopy ◽  
Light Microscopy ◽  
Ultrastructural Level ◽  
Electron Microscopic ◽  
Biological Interest ◽  
Investigative Approach ◽  
Immunologic Activity ◽  
Dense Label

Immunocytochemistry is a powerful investigative approach in which one of the most exacting examples of specificity, that of the reaction of an antibody with its antigen, isused to localize tissue and cell specific molecules in situ. Following the introduction of fluorescent labeled antibodies in T950, a large number of molecules of biological interest had been studied with light microscopy, especially antigens involved in the pathogenesis of some diseases. However, with advances in electron microscopy, newer methods were needed which could reveal these reactions at the ultrastructural level. An electron dense label that could be coupled to an antibody without the loss of immunologic activity was desired.

Electron Microscopy of Mycoplasma Pneumoniae

1971 ◽  
Vol 29 ◽  
pp. 258-259 ◽  
Author(s):  
E. S. Boatman ◽  
G. E. Kenny
Keyword(s):  
Electron Microscopy ◽  
Light Microscopy ◽  
Growth Phase ◽  
Hela Cells ◽  
Sulfonic Acid ◽  
Gamma Globulin ◽  
Horse Serum ◽  
Morphological Aspects ◽  
The Family

Information concerning the morphology and replication of organism of the family Mycoplasmataceae remains, despite over 70 years of study, highly controversial. Due to their small size observations by light microscopy have not been rewarding. Furthermore, not only are these organisms extremely pleomorphic but their morphology also changes according to growth phase. This study deals with the morphological aspects of M. pneumoniae strain 3546 in relation to growth, interaction with HeLa cells and possible mechanisms of replication.The organisms were grown aerobically at 37°C in a soy peptone yeast dialysate medium supplemented with 12% gamma-globulin free horse serum. The medium was buffered at pH 7.3 with TES [N-tris (hyroxymethyl) methyl-2-aminoethane sulfonic acid] at 10mM concentration. The inoculum, an actively growing culture, was filtered through a 0.5 μm polycarbonate “nuclepore” filter to prevent transfer of all but the smallest aggregates. Growth was assessed at specific periods by colony counts and 800 ml samples of organisms were fixed in situ with 2.5% glutaraldehyde for 3 hrs. at 4°C. Washed cells for sectioning were post-fixed in 0.8% OSO4 in veronal-acetate buffer pH 6.1 for 1 hr. at 21°C. HeLa cells were infected with a filtered inoculum of M. pneumoniae and incubated for 9 days in Leighton tubes with coverslips. The cells were then removed and processed for electron microscopy.

Ectodesmata as Revealed By Freeze-Etch Preparations of Plant Epidermal Cell Walls

1973 ◽  
Vol 31 ◽  
pp. 468-469
Author(s):  
N.C. Lyon ◽  
W. C. Mueller
Keyword(s):  
Electron Microscopy ◽  
Acetic Acid ◽  
Nitric Acid ◽  
Oxalic Acid ◽  
Light Microscopy ◽  
Epidermal Cell ◽  
Cell Walls ◽  
Plant Viruses ◽  
Plant Leaves ◽  
Thin Sections

Schumacher and Halbsguth first demonstrated ectodesmata as pores or channels in the epidermal cell walls in haustoria of Cuscuta odorata L. by light microscopy in tissues fixed in a sublimate fixative (30% ethyl alcohol, 30 ml:glacial acetic acid, 10 ml: 65% nitric acid, 1 ml: 40% formaldehyde, 5 ml: oxalic acid, 2 g: mecuric chloride to saturation 2-3 g). Other workers have published electron micrographs of structures transversing the outer epidermal cell in thin sections of plant leaves that have been interpreted as ectodesmata. Such structures are evident following treatment with Hg++ or Ag+ salts and are only rarely observed by electron microscopy. If ectodesmata exist without such treatment, and are not artefacts, they would afford natural pathways of entry for applied foliar solutions and plant viruses.

A Method for Electron Microscopic Study of Tissue Culture Cell Monolayers Grown in Tubes

1967 ◽  
Vol 25 ◽  
pp. 132-133
Author(s):  
R. Stephens ◽  
G. Schidlovsky ◽  
S. Kuzmic ◽  
P. Gaudreau
Keyword(s):  
Electron Microscopy ◽  
Tissue Culture ◽  
Light Microscopy ◽  
Cell Sheet ◽  
Culture Cell ◽  
Tissue Culture Cell ◽  
Electron Microscopic ◽  
Cell Sheets ◽  
Morphological Alterations ◽  
Culture Cells

The usual method of scraping or trypsinization to detach tissue culture cell sheets from their glass substrate for further pelletization and processing for electron microscopy introduces objectionable morphological alterations. It is also impossible under these conditions to study a particular area or individual cell which have been preselected by light microscopy in the living state.Several schemes which obviate centrifugation and allow the embedding of nondetached tissue culture cells have been proposed. However, they all preserve only a small part of the cell sheet and make use of inverted gelatin capsules which are in this case difficult to handle.We have evolved and used over a period of several years a technique which allows the embedding of a complete cell sheet growing at the inner surface of a tissue culture roller tube. Observation of the same cell by light microscopy in the living and embedded states followed by electron microscopy is performed conveniently.

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