Viability cell counting

Trypan Blue exclusion and viable cell counts Cell viability by MTT... 

Immunopathology Hematology (blood cell count), weight of spleen, thymus and liver, histology of spleen, thymus, and lymph nodes Histopathology, cell viability, lysosomal membrane stability... 

Count the cells by trypan blue exclusion test in a Biirker-Turk cell counting chamber and calculate the correct dilution. Note The cell viability should be >90 %. 

The use of trypsin is common to detach cells from adherent cultures however, in this case it is not recommended. MCF-7 cells treated with trypsin tend to form clumps, which are difficult to breakup. The use of Accutase yields a mostly single-cell suspension that will not form clumps even after it has been washed with labeling buffer. Total cell count and viability assays will also be easier for the researcher to carry out. 

We have found that turbidity measurements can be used as a routine test to keep track of the yeast cell population. Fermentations routinely were examined microscopically for cell count and viability. We use a 1% methylene blue stain to differentiate viable and dead yeast cells. The dead cells take the stain. 

The cells in the biological fluids are stained with trypan blue for determination of viability and with crystal violet containing citric acid (0.5 gm crystal violet and 10.5 grams citric acid in 500 mL dH20) for cell counts. 

To ensure a constant and comparable viability cells are counted and cell viability is determined with trypan... 

Figure 4 A photograph of the Selec H robot built by The Technology Partnership, Cambridge, UK. This robot maintains between 1 and 50 cell lines, performs passaging, cell counting and viability measurements, and direct plating into microtitre plates for bioassay...

Automation of cell counting (total cells) is possible with electronic counters, especially for non-clumping single suspension cells. A method that is widely used for total cell numbers is counting cell nuclei after dissolving the cytoplasm. This is particularly useful for large clumps of cells, where cells are inaccessible (e.g. in matrices) or where cells are difficult to trypsinize off substrates (e.g. microcarriers). If determination of viability is the prime consideration, then the most accurate method is to count the cells that have the ability to divide by the mitotic index method. 

In order to ensure that cell cultures have reached the optimum level of growth before routine subculture or freezing, it is helpful to obtain an accurate cell count and a measure of the percentage viability of the cell population. 

Remove 1 ml of cell suspension into a disposable culture tube for a cell count. Determine the cell concentration of the suspension by direct counting and monitor the viability using the Trypan blue exclusion test. 

The use of an infrared sensor (Aquasant Messtechnik AG, Bubendorf, Switzerland) may be a solution for many apphcations, particularly in combination with a control of viability (Merten et al, 1987). Other alternatives are sensors measuring conductance/capacitance (ABER Instruments, Cefnllan, Aberystwyth, UK) and software sensors (Pelletier et al, 1994). For further reading see De Gouys et al (1996). A recently pubhshed method based on real-time imaging opens up new possibihties by real cell counting (Ozturk et al, 1997). 

Figure 12.3. Effects of HT (41°C) or USMH or verapamil (Ver) on cytotoxicity enhancement of doxorubicin (DOX) in the parent or MDR variants of human MV522 and KB cell lines. Data are expressed as % inhibition calculated by the following formula % inhibition = [1 — (counts of viable drug-exposed cells/counts of viable non-drug-exposed cells)] x 100. Cell viability was determined by hemocytometry technique after trypan blue staining. All experiments were carried out in triplicates. P < 0.05 compared to control values.

FIGURE 1 Yeast cell growth, viability, sugar consumption, and ethanol production patterns of a typical fermentation. In a synthetic grape juice medium. Triple M (Spiropoulos et al., 2000) was used and inoculated with a commercial strain of 5. cerevisiae. Glucose and fructose concentrations were determined by enzymatic assay, viable cell counts by plating on YPD medium, and cell mass by absorbance at 580 nm. 

A 20X concentration was obtained in a six hour processing time with an average flux of 15 GFD. Titrations of the initial and final viable cell counts illustrate the efficient recovery of the cells in this process with no loss of viability. 

Saturation density assay Untransformed cells, which are still contact inhibited, will grow in a monolayer, whereas transformed cells will grow on top of each other and thus can be grown to a higher density which can be measured by the different viability assays or by a higher cell count. 

After 20-24 h of incubation, count the cells and measure viability using trypan blue. At this stage, cell growth should be arrested (total cell count 3 X 10° ceUs/mL) and the cell diameter increased by 20-25% compared to control cells, but the viabihty should remain... 

After 48 h total incubation, count the cells and measure the viability. At this point, the viabihty will be lowered (usually 60—80% for chemokines and 50—70% for receptors) and the total cell count is typically 3 x 10 cells/mL. 

Brewing yeast cell count/viability/vitality methods... 

Exposure of an immortalised rat type II alveolar epithelial cell line to 10 fM CdClj for 24 h caused a significant decrease in viability (Timblin et al. 1998). After 48 h of exposure to 1,5, or 10 /iM CdCU a dose-dependent decrease in viability was observed at dl concentrations as determined by trypan blue exclusion and cell counting. Significant increases in reduced and oxidised glutathione levels were observed at 24 and 48 h in rat lung epithelial cells exposed to CdC. Hart et al. (1999) observed a maximum level of apoptosis (5-fold higher than control) in cultures exposed for 48 h to 20/ M... 

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