Viable Cell Counting

Isotonic saline (0.7% w/v) In approximately 950 mL of distilled water, dissolve 7 g NaCL and bring to 1 L volume with distilled water. 

Before beginning, it may be advisable to review earlier sections (C.2) for basic protocol. 

Depending on the stage of yeast growth, prepare the appropriate dilutions using 9-mL isotonic saline dilution blanks. For example, in monitoring starter tanks, initial lO -lO dilutions may be required to obtain countable numbers, whereas post-fermentation counts may be made directly without prior dilution. 

Vortex each diluted sample before going to the next dilution in the series. 

After vortexing the final dilution in the series, place the cover-slip (provided with kit) over the counting grid of the hemocytometer. 

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Fig. 3. Viable cell count of adherent BHK cells as a function of temperature and shear stress...

Others Molecular probes— stain all, vital stain Vital stains particularly useful for total viable cell counts 122... 

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

Perform a viable cell count using trypan blue stain exclusion and adjust the viable cell concentration to the desired number per milliliter with CMF-HBSS. 

Batch culture experiments were performed to measure the rate of glucose consumption. Once the seed yeast had been incubated for exactly 48 h, the serum vials were each injected with 0.2 mL of freshly vortexed yeast. The moment of inoculation became time zero (f = 0) for the experiment. Viable cell counts taken at time zero showed the initial cell concentration to be on the order of 100,000 colony-forming units/mL. Samples were taken every 2 h over the next 12 h, except for the first 4 hours, and analyzed for glucose. 

Because the cells are viewed directly, this method enables a judgement to be made about the quality of the cell suspension, i.e. clumped or broken cells present. By diluting the cell suspension with a solution of a vital dye, e.g. trypan blue at a final concentration of 0.2 g/1 PBS-A (Appendix 1), and counting only the unstained cells a measure of the viable cell count is obtained. (N.B. Trypan blue is toxic and should not be allowed to come in contact with the skin.)... 

Total cell count = viable cell count + dead cell count... 

Connect a glass bell-end attachment to the multiplate-end connection. Pressurize the harvest vessel and, via the glass bell-end, remove a small volume of cell suspension for a viable cell count. An MP unit seeded with a minimum of 70 X 10 cells will yield approximately 0.8-1.6 x 10 human diploid cells. For operations involving viral infection instead of trypsinization, the following steps should be considered. 

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. 

Water sorption, viable cell count of Rhizopus japonicum, relative mobile protons and deuterons and T2 relaxation times for freeze-dried xanthan and locust bean gum gels. (Source From Vittadini, E., Dickinson, L.C., and Chinachoti, P. NMR water mobility in xanthan and locust bean gum mixtures possible explanation of microbial response. Curb. Poly., 49,261,2002. With permission.)... 

With some biocide treatments there is a simple direct relationship between ATP and viable cell counts. However, where the mechanism of biocide action is related to influences on cell bioenergetics, or the concentration range employed is biostatic and affects cell metabolism, then ATP results may not follow the same pattern as microbial counts. Under these circumstances low ATP results at relatively high microbial levels might be expected but it is also possible that ATP levels increase over a period of time. 

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. 

The death phase (involving exponential/logarith-mic decline) begins when cell deaths exceed births (i.e. the viable-cell count declines). 

Fig. 4 Viable cell count during the fermentation of bacterial cellulose on a coconut juice and b pineapple juice...

Sample from brewery no. Total bacterial count (living and dead) per 10 yeast cells Viable cell count (per 10 yeast cells) ... 

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