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In the study of microbiology, there are numerous occasions when it is necessary to

either estimate or determine the number of bacterial cells in a broth culture or


liquid medium. Determination of cell numbers can be accomplished by a number
of direct or indirect methods. The methods include standard plate counts,
turbidimetric measurements, visual comparison of turbidity with a known standard,
direct microscopic counts, cell mass determination, and measurement of cellular
activity. There are 3 methods of bacterial enumeration: viable counts, total cell
count by using Haemocytometer, and turbidimetric measurement

A serial dilution is a series of sequential dilutions used to reduce a dense culture


of cells to a more usable concentration. Each dilution will reduce the concentration of bacteria
by a specific amount. Serial dilution method is carried out to estimate the concentration (number
of microbes) of an unknown sample by counting the number of colonies cultured from serial
dilutions of the sample, and then back track the measured counts to the unknown
concentration. (Ben-David and Davidson, 2014). 1 mL of culture from the original inoculum is
added into 9 mL of broth in a test tube labelled 10-1, which it will now contain 1/10 the number
of cells present in the original sample. Then, another 1 mL of dilution in the 10-1 test tube is the
transferred into the next test tube labelled 10-2, a dilution factor of 10-2. The procedure of serial
dilution is repeated until the last tube contains 1/100000000 of the original bacterial culture, with
dilution factor of 10-8 is obtained. (Keler et al., 2010)

A viable cell count is essential to evaluate the kinetics of cell growth and is crucial for the study
of eukaryotic cells for different purposes. The suspension is either spread onto the surface of agar
plates (spread plate method), or is mixed with molten agar, poured into plates, and allowed to
solidify (pour plate method). 0.1 mL of the diluted culture with dilution factor 106, 107 and 106
are transferred to each agar plate respectively. Streaking technique using bent glass spreader is
used to distribute the bacteria on the surface of agar plate to ensure evenly bacterial growth.
(Cadena-Herrera et al., 2015). The number of colonies on a dilution plate showing a range
between between 30 and 300 colonies is considered statistically effective. Fewer than 30
colonies makes a poorer statistical precision while greater than 300 colonies often
results in overlapping colonies and imprecision in the count. (Cadena-Herrera et al., 2015)
(Sieuwerts et al., 2008)
Ben-David, A. and Davidson, C. E. (2014) Estimation method for serial dilution experiments,
Journal of Microbiological Methods, 107, pp. 214221. doi: 10.1016/j.mimet.2014.08.023.
Cadena-Herrera, D. et al. (2015) Validation of three viable-cell counting methods: Manual, semi-
automated, and automated, Biotechnology Reports, 7, pp. 916. doi:
10.1016/j.btre.2015.04.004.
Keler, C. et al. (2010) Serial Dilution Simulation Lab, The American Biology Teacher, 72(5), pp.
305307. doi: 10.1525/abt.2010.72.5.9.
Sieuwerts, S. et al. (2008) A simple and fast method for determining colony forming units,
Letters in Applied Microbiology, 47(4), pp. 275278. doi: 10.1111/j.1472-765X.2008.02417.x.