SINGLE UV EXCITATION OF HOECHST 33342 AND PROPIDIUM IODIDE FOR VIABILITY ASSESSMENT OF RHESUS MONKEY SPERMATOZOA USING FLOW CYTOMETRY

2005 ◽  
Vol 51 (5) ◽  
pp. 371-383 ◽  
Author(s):  
K. Cai ◽  
J. Yang ◽  
M. Guan ◽  
W. Ji ◽  
Y. Li ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Rhesus Monkey ◽  
Propidium Iodide ◽  
Viability Assessment ◽  
Hoechst 33342 ◽  
Uv Excitation

scholarly journals Chromosome doubling in Cattleya tigrina A. Rich

2019 ◽  
Vol 15 (11) ◽  
Author(s):  
Thays Saynara Alves Menezes-Sá ◽  
Maria de Fátima Arrigoni-Blank ◽  
Andréa Santos da Costa ◽  
Janay De Almeida Santos-Serejo ◽  
Arie Fitzgerald Blank ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Propidium Iodide ◽  
Chromosome Doubling ◽  
Flow Cytometric Analysis ◽  
Leaf Tissue ◽  
Stomatal Index ◽  
Chromosome Duplication ◽  
Flow Cytometric ◽  
Young Leaves ◽  
Commercial Value

Chromosome doubling induction in orchids may benefit their production for resulting in flowers of higher commercial value, larger size and higher content of substances that intensify the color and fragrance when compared with diploid orchids. This work aimed to induce and confirm artificial polyploidization, using flow cytometry and stomatal analysis. Explants were treated with colchicine at concentrations of 0, 2.5, 7.5, and 12.5 mM, for 24 and 48 hours and with oryzalin, at concentrations of 0, 10, 30, and 50 μM, for three and six days. For the flow cytometric analysis, a sample of leaf tissue was removed from each plant, crushed to release the nuclei and stained with propidium iodide. In addition to flow cytometry, the ploidy of the antimitotic treated plants was evaluated by stomata analysis. Young leaves were used where the density, functionality and stomatal index were evaluated. Colchicine provided induction of satisfactory polyploidy in C. tigrina at all concentrations and times of exposure, obtaining a greater number of polyploid individuals in the concentration of 12.5 mM for 48 hours. Oryzalin did not induce chromosome duplication at the tested concentrations.

Flow cytometry and cell sorting for yeast viability assessment and cell selection

Yeast ◽  
1998 ◽  
Vol 14 (2) ◽  
pp. 147-160 ◽  
Author(s):  
Daniel Deere ◽  
Jian Shen ◽  
Graham Vesey ◽  
Philip Bell ◽  
Peter Bissinger ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Sorting ◽  
Cell Selection ◽  
Viability Assessment ◽  
Yeast Viability

scholarly journals Data-Driven Compensation for Flow Cytometry of Solid Tissues

2011 ◽  
Vol 2011 ◽  
pp. 1-11 ◽  
Author(s):  
Nickolaas Maria van Rodijnen ◽  
Math Pieters ◽  
Sjack Hoop ◽  
Marius Nap
Keyword(s):  
Flow Cytometry ◽  
Propidium Iodide ◽  
Dna Content ◽  
Flow Cytometer ◽  
Data Driven ◽  
Color Flow ◽  
Operator Experience ◽  
Solid Tissues

Propidium Iodide is a fluorochrome that is used to measure the DNA content of individual cells, taken from solid tissues, with a flow cytometer. Compensation for spectral cross-over of this fluorochrome still leads to compensation results that are depending on operator experience. We present a data-driven compensation (DDC) algorithm that is designed to automatically compensate combined DNA phenotype flow cytometry acquisitions. The generated compensation values of the DDC algorithm are validated by comparison with manually determined compensation values. The results show that (1) compensation of two-color flow cytometry leads to comparable results using either manual compensation or the DDC method; (2) DDC can calculate sample-specific compensation trace lines; (3) the effects of two different approaches to calculate compensation values can be visualized within one sample. We conclude that the DDC algorithm contributes to the standardization of compensation for spectral cross-over in flow cytometry of solid tissues.

461 Prostate stem cell isolation using Hoechst 33342 flow cytometry

2004 ◽  
Vol 3 (2) ◽  
pp. 118
Author(s):  
P. Gilmore ◽  
R. Bhatt ◽  
C. Hart ◽  
V. Ramani ◽  
N. George ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Stem Cell ◽  
Cell Isolation ◽  
Hoechst 33342 ◽  
Stem Cell Isolation

scholarly journals Intermittent blood flow in the KHT sarcoma--flow cytometry studies using Hoechst 33342

1990 ◽  
Vol 62 (2) ◽  
pp. 195-200 ◽  
Author(s):  
AI Minchinton ◽  
RE Durand ◽  
DJ Chaplin
Keyword(s):  
Flow Cytometry ◽  
Blood Flow ◽  
Hoechst 33342

scholarly journals Use of Formalin-Fixed, Propidium Iodide-Stained Human Leukocytes as a Standard for Enumerating CD4+ T Lymphocytes in a Single-Platform Assay

2001 ◽  
Vol 8 (2) ◽  
pp. 397-401 ◽  
Author(s):  
G. M. Harrison ◽  
A. J. Bennett ◽  
M. Moody ◽  
G. F. Read ◽  
P. E. Williams
Keyword(s):  
Flow Cytometry ◽  
Blood Cell ◽  
Propidium Iodide ◽  
Internal Standard ◽  
Inexpensive Method ◽  
Normal Cells ◽  
Blood Samples ◽  
Cell Enumeration ◽  
Whole Blood Samples ◽  
Formalin Fixed

ABSTRACT A new, inexpensive method is described that enables lymphocytes to be enumerated very precisely. Normal leukocytes were simultaneously stained and fixed with a propidium iodide-paraformaldehyde solution. The preparation obtained (CellBeads) was used as an internal standard for cell enumeration by flow cytometry and was stable at 4°C for at least 60 days. Unlike synthetic beads, the CellBeads behaved similarly to normal cells during red blood cell lysis and cell washing procedures. When known numbers of CellBeads were added to whole-blood samples and the numbers of CellBeads and lymphocytes were determined, highly reproducible and accurate enumerations were obtained—far more so than when synthetic beads were used. This inexpensive method is suitable for routine use.

scholarly journals Measurement of cell-cycle phase-specific cell death using Hoechst 33342 and propidium iodide: preservation by ethanol fixation.

1988 ◽  
Vol 36 (9) ◽  
pp. 1147-1152 ◽  
Author(s):  
G Ciancio ◽  
A Pollack ◽  
M A Taupier ◽  
N L Block ◽  
G L Irvin
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Propidium Iodide ◽  
Cell Cycle Phase ◽  
Cytotoxic Agents ◽  
Cycle Phase ◽  
Specific Cell ◽  
Dead Cell ◽  
Hoechst 33342 ◽  
The Dead

We developed a rapid technique for preservation of Hoechst 33342/propidium iodide-stained cells, using ethanol as a fixative. Combined staining with these dyes makes possible analysis of cell-cycle phase-specific cell death. The technique relies on exclusion of propidium iodide from the viable cells, whereas Hoechst stains all of the cells. The bivariate histograms resulting from the flow cytometric analysis contain the equivalent of two single-parameter DNA histograms, one of the living and the other of the dead cell population. Preservation of staining involved addition of 25% ethanol in PBS after propidium iodide staining and before Hoechst staining. The separation between the living and the dead cell populations was maintained for over 3 days at 4 degrees C. This technique will be valuable for quantitative evaluation of the cell-cycle phase-specific effects of cytostatic or cytotoxic agents, particularly in situations where a lag period between staining and analysis is unavoidable.

Pannexin channels increase propidium iodide permeability in frozen–thawed dog spermatozoa

2017 ◽  
Vol 29 (11) ◽  
pp. 2269 ◽  
Author(s):  
J. L. Torres ◽  
J. Palomino ◽  
R. D. Moreno ◽  
M. De los Reyes
Keyword(s):  
Flow Cytometry ◽  
Fluorescence Microscopy ◽  
Analysis Of Variance ◽  
Propidium Iodide ◽  
Membrane Channels ◽  
Flow Cytometry Data ◽  
Viable Cells ◽  
Head Segment ◽  
Relationship Of ◽  
The Relationship

Pannexins (Panx) are proteins that form functional single membrane channels, but they have not yet been described in dogs. The aim of the present study was to detect Panx1, Panx2 and Panx3 in frozen–thawed dog spermatozoa using flow cytometry and immunofluorescence analyses, evaluating the relationship of these proteins with propidium iodide (PI) in frozen–thawed spermatozoa. Fresh and frozen–thawed dog spermatozoa from eight dogs were preincubated with 3 μM PI with or without 15 μM carbenoxolone (CBX) or 1 mM probenecid (PBD), two Panx channel inhibitors, and then incubated with rabbit anti-Panx1, anti-Panx2 and anti-Panx3 antibodies (1 : 200). Panx immunolocalisation was assessed by fluorescence microscopy. Flow cytometry data were evaluated by analysis of variance. All three Panx proteins were found in dog spermatozoa: Panx1 was mostly localised to the acrosomal and equatorial segment, Panx2 was found in the posterior region of the head and tail and Panx3 was localised to the equatorial and posterior head segment. The percentage of PI-positive cells determined by flow cytometry was reduced (P < 0.05) in the presence of Panx inhibitors. These results show that Panx proteins are present in dog spermatozoa and increase PI permeability in frozen–thawed dog sperm, suggesting that the percentage of PI-positive spermatozoa used as an indicator of non-viable cells may lead to overestimation of non-viable cells.

110 COMPARISON OF ANDROMED®, BIOXCELL®, AND BOTU-BOV® EXTENDERS FOR CRYOPRESERVATION OF BULL SEXED SEMEN

2007 ◽  
Vol 19 (1) ◽  
pp. 172 ◽  
Author(s):  
M. Q. Braga ◽  
R. V. R. Franco ◽  
L. F. Rodrigues ◽  
G. Galeli ◽  
K. M. Oliveira ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Semen Quality ◽  
Membrane Damage ◽  
Optical Microscope ◽  
Hoechst 33342 ◽  
Tukey Test ◽  
Progressive Motility ◽  
Significant Difference ◽  
Sexed Semen ◽  
Fort Collins

Sexing semen has become a worldwide technology now available in many countries through the use of flow cytometry for sexing mammal sperms (Johnson and Welch 1999 Theriogenology 52, 1323–1341). Because straws containing sexed semen have a low concentration, any condition that either improves or decreases freezing capabilities will considerably change semen quality. During cryopreservation, spermatozoa have been described as undergoing many changes that lead to membrane damage, which may result in decreased fertility (Watson 2000 Reprod. Fertil. Dev. 6 (Suppl 1), 481). Since many cryoprotectants are available on the market, the objective of the present study was to compare 3 different extenders for freezing sex-sorted semen. For this study, 25 ejaculates were collected from 8 bulls of different breeds, diluted, then dyed with Hoechst 33342 (Schenk et al. 1999 Theriogenology 52, 1375–1391), and sexed by flow cytometry (SX MoFlo®; DaKoCytomation, Inc., Fort Collins, CO, USA). After being cooled at 4°C for 1 h and 30 min, the sexed semen was centrifuged and diluted in AndroMed® (Minitüb, Tiefenbach, Germany), Bioxcell® (IMV, Aigle, France), or Botu-Bov® (Biotech Botucatu, Ltda., Sao Paulo, Brazil); the semen was packaged at 3 million total sperm in 0.25-mL straws and frozen in an automatic freezer (Digit cool 5300® IMV). To evaluate the freezing quality, the straws were thawed and incubated at 35°C for 15 min. The progressive motility was observed through an optical microscope (Coleman 200T). The statistical analyses were done using the SAS program (SAS Institute, Inc., Cary, NC, USA) and the Tukey test (P ≤ 0.05). Results show that there was no statistical difference between Bioxcell and AndroMed extenders (P ≤ 0.05). However, Botu-Bov extender showed a significant difference when compared with Bioxcell and AndroMed (see Table 1). It is also important to point out that 40% of the samples frozen with AndroMed showed non-aligned movement. Even though few ejaculates were used for this study, preliminary results showed that Bioxcell seemed to be the most suitable for freezing bull sexed semen. Table 1. Percentage of progressively motile spermatozoa after thawing

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