Evaluation of a fluorescent method (fluorescein diacetate and ethidium bromide solution) in the study of the viability Cryptococcus neoformans strains

1986 ◽  
Vol 96 (2) ◽  
pp. 91-96 ◽  
Author(s):  
B. Correa ◽  
A. Purchio ◽  
C. R. Paula ◽  
W. Gambale
Keyword(s):  
Cryptococcus Neoformans ◽  
Ethidium Bromide ◽  
Fluorescein Diacetate ◽  
Ethidium Bromide Solution ◽  
Fluorescent Method ◽  
Bromide Solution

The Use of the Vitality and Chemotaxis of Human Polymorphonuclear Leukocytes for the In Vitro Estimation of the Acute Toxicity of the First Ten Chemicals from the MEIC List

1993 ◽  
Vol 21 (1) ◽  
pp. 73-80
Author(s):  
Matteo Valentino ◽  
Francesca Monaco ◽  
Maria Antonietta Pizzichini ◽  
Mario Governa
Keyword(s):  
Ethidium Bromide ◽  
Polymorphonuclear Leukocytes ◽  
Rank Correlation ◽  
Fluorescein Diacetate ◽  
Live Cells ◽  
In Vitro Toxicity ◽  
Ic50 Values ◽  
Acute Cytotoxicity ◽  
Human Polymorphonuclear Leukocytes

The acute cytotoxicity of the first ten MEIC chemicals has been estimated by others in various cell lines. In the present investigation, isolated human polymorphonuclear leukocytes (PMN) from ten healthy non-smoking laboratory personnel were used to assess in vitro toxicity of the same chemicals. The cells were treated with different concentrations of the respective chemicals for three hours and their vitality and chemotaxis were tested. Vitality was measured by fluorescence microscopy after the addition of fluorescein diacetate and ethidium bromide. Living cells which took up and hydrolysed fluorescein diacetate, and dead cells, stained by ethidium bromide, were counted and the percentage of live cells was calculated. Locomotion stimulated by the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (F-MLP), was measured in blind-well Boyden chambers and a chemotactic index was calculated. The results were mathematically transformed to produce a linear curve, and then fitted by the linear least squares procedure, from which LC50 and IC50 values were obtained by interpolation. All the chemicals decreased the vitality and inhibited the chemotaxis of the PMN. Obviously the chemotactic test was more sensitive than the vitality one. A correlation (r = 0.933) was found between vitality and chemotaxis inhibition. Spearman rank correlation analysis revealed significant correlations between our results and those from in vitro experiments conducted in other laboratories, as well as with data concerning mouse, rat and human lethal doses.

scholarly journals Remyelination in experimentally demyelinated connexin 32 KnockOut mice

2009 ◽  
Vol 67 (2b) ◽  
pp. 488-493
Author(s):  
Adriano Tony Ramos ◽  
Paulo César Maiorka ◽  
Maria Lúcia Zaidan Dagli ◽  
Fernando Yutaka Moniwa Hosomi ◽  
Kalan Bastos Violin ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Connexin 32 ◽  
Ethidium Bromide Solution ◽  
Normal Pattern ◽  
Myelin Sheaths ◽  
Transmission Electron ◽  
Bromide Solution ◽  
Sciatic Nerves ◽  
Electron Microscopical

The aim of this study was to evaluate the role of connexin 32 (Cx 32) during remyelination of the peripheral nervous system, through a local injection of either 0,1% ethidium bromide solution or saline in the sciatic nerve of Cx 32 knockout mice. Euthanasia was performed ranging from 1, 2, 3, 7, 15, 21 to 30 days after injection. Histochemical, immunohistochemical, immunofluorescence and transmission electron microscopical techniques were used to analyze the development of the lesions. Within the sciatic nerves, Schwann cells initially showed signs of intoxication and rejected their sheaths; after seven days, some thin newly formed myelin sheaths with uneven compactness and redundant loops (tomacula) were conspicuous. We concluded that the regeneration of lost myelin sheaths within the PNS followed the pattern already reported for this model in other laboratory species. Therefore, these results suggest that absence of Cx 32 did not interfere with the normal pattern of remyelination in this model in young mice.

scholarly journals In vitro action of some disinfectants on Paracoccidioides brasiliensis yeast forms

1991 ◽  
Vol 33 (1) ◽  
pp. 37-43
Author(s):  
M. A. Shikanai-Yasuda ◽  
R. T. Taguchi ◽  
M. K. Sato ◽  
N. T. Melo ◽  
C. M. Assis ◽  
...  
Keyword(s):  
Ethidium Bromide ◽  
Metabolic Activity ◽  
Room Temperature ◽  
Paracoccidioides Brasiliensis ◽  
Fluorescein Diacetate ◽  
Liquid Media ◽  
Fungicidal Action ◽  
Viable Cells ◽  
Treatment Culture

The fungicidal action of sodium hypochlorite (0.3, 1, 2.5, 5 and 10%); formaldehyde (2, 5, and 10%); and ethyl alcohol (70%) on yeast forms of Paracoccidioides brasiliensis Pb 18 and a newly-isolated Goiana strain was described. Contact between the fungus and the disinfectants was maintained for 1, 2, 24, 48 and 72 hours at room temperature. Viability was evaluated by the fluorescein diacetate-ethidium bromide treatment, culture in solid and liquid media (36ºC and 26ºC); yeast to mycelial germination at room temperature; and radiometric study of metabolic activity. All concentrations of disinfectants were found to be effective in inactivating Pb 18 and Goiana strains, except for the 1-hour contact with 2% formaldehyde, in which fluorescein diacetate-ethidium bromide treatment was found to reveal 40 and 27% of viable cells, respectively. The yeast to mycelial germination method was considered to reveal faster and similar results as compared to culture in solid and liquid media.

Establishment of Fluorescein Diacetate and Ethidium Bromide (FDAEB) Assay for Quality Assessment of Isolated Islets

2000 ◽  
Vol 9 (5) ◽  
pp. 681-686 ◽  
Author(s):  
Masaaki Miyamoto ◽  
Yoshihiko Morimoto ◽  
Yuka Nozawa ◽  
A. N. Balamurugan ◽  
Baoyou Xu ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Quality Assessment ◽  
Islet Transplantation ◽  
Ethidium Bromide ◽  
Isolated Islets ◽  
Fluorescein Diacetate ◽  
Live Cells ◽  
Viability Assay ◽  
Clinical Islet Transplantation ◽  
Light Excitation

One of the most important factors in clinical islet transplantation is isolation of a great number of islets with good viability. According to viability assessments of isolated islets, the static incubation test and the perifusion test of islets, which are used retrospectively, take much time and need various apparatus. But viability assessments of isolated islets for clinical islet transplantation require a simple, rapid, sensitive, and prospective method. We have developed a microfluorometric viability assay for isolated human, porcine, and dog islets of pancreata using fluorescein diacetate and ethidium bromide (FDAEB). Fluorescein diacetate (FDA) causes live cells to fluoresce green under blue light excitation (490 nm) and ethidium bromide (EB) causes dead cells to fluoresce red. In this study, we investigated the applicability of FDAEB staining to quality assessment of isolated islets for clinical use by correlation with the counting method with insulin secretion of islets. Discrimination of living from dead islets by insulin secretion correlated well with viability as determined by FDAEB staining. The proportion of living islets within isolated canine islets, as measured by microfluorometric counting, was found to correlate highly significantly on low-temperature (24°C) culture (R = 0.831, p < 0.001) and on 37°C culture (R = 0.553, p < 0.05) with the insulin contents of the same islets. Therefore, it is possible to differentiate degrees of viability, and a scoring system is described for this purpose. The FDAEB assay prospectively and easily provides a rapid, accurate, and objective measurement of the proportion of living cells and dead cells in isolated islets for clinical islet transplantation.

Fluorescein diacetate and ethidium bromide staining to determine the viability ofMycobacterium smegmatisandEscherichia coli

1991 ◽  
Vol 62 (3) ◽  
Author(s):  
V. JAYAPAL ◽  
K. M. SHARMILA ◽  
G. SELVIBAI ◽  
S. P. THYAGARAJAN ◽  
N. SHANMUGASUNDARAM ◽  
...  
Keyword(s):  
Ethidium Bromide ◽  
Fluorescein Diacetate ◽  
Ethidium Bromide Staining

Properties of Isolated Mitochondria of Agaricus Bisporus: Their Relationship to Dormancy and the Germination of Spores

1973 ◽  
Vol 31 ◽  
pp. 458-459
Author(s):  
K. S. McCarty ◽  
R. F. Weave ◽  
L. Kemper ◽  
F. S. Vogel
Keyword(s):  
Mitochondrial Dna ◽  
Ethidium Bromide ◽  
Microscopic Examination ◽  
Agaricus Bisporus ◽  
Osmotic Shock ◽  
Electron Microscopic Examination ◽  
Electron Microscopic ◽  
Isolated Mitochondria ◽  
Dna Strands ◽  
Cytoplasmic Ribosomes

During the prodromal stages of sporulation in the Basidiomycete, Agaricus bisporus, mitochondria accumulate in the basidial cells, zygotes, in the gill tissues prior to entry of these mitochondria, together with two haploid nuclei and cytoplasmic ribosomes, into the exospores. The mitochondria contain prominent loci of DNA [Fig. 1]. A modified Kleinschmidt spread technique1 has been used to evaluate the DNA strands from purified whole mitochondria released by osmotic shock, mitochondrial DNA purified on CsCl gradients [density = 1.698 gms/cc], and DNA purified on ethidium bromide CsCl gradients. The DNA appeared as linear strands up to 25 u in length and circular forms 2.2-5.2 u in circumference. In specimens prepared by osmotic shock, many strands of DNA are apparently attached to membrane fragments [Fig. 2]. When mitochondria were ruptured in hypotonic sucrose and then fixed in glutaraldehyde, the ribosomes were released for electron microscopic examination.

Deformation of Yeast Plasma Membrane by Ethidium Bromide

1988 ◽  
Vol 46 ◽  
pp. 254-255
Author(s):  
E. Keyhani
Keyword(s):  
Plasma Membrane ◽  
Thin Section ◽  
Ethidium Bromide ◽  
Freeze Fracture ◽  
Mutagenic Effect ◽  
Yeast Cells ◽  
Hydrophobic Region ◽  
Thin Section Electron Microscopy ◽  
Section Electron ◽  
Deep Pits

The mutagenic effect of ethidium bromide on the mitochondrial DNA is well established. Using thin section electron microscopy, it was shown that when yeast cells were grown in the presence of ethidium bromide, besides alterations in the mitochondria, the plasma membrane also showed alterations consisting of 75 to 110 nm-deep pits. Furthermore, ethidium bromide induced an increase in the length and number of endoplasmic reticulum and in the number of intracytoplasmic vesicles.Freeze-fracture, by splitting the hydrophobic region of the membrane, allows the visualization of the surface view of the membrane, and consequently, any alteration induced by ethidium bromide on the membrane can be better examined by this method than by the thin section method.Yeast cells, Candida utilis. were grown in the presence of 35 μM ethidium bromide. Cells were harvested and freeze-fractured according to the procedure previously described.

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