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Instructions on how to prepare dilutions and concentrations of microbiological samples, including liquid and solid samples. It explains the concept of dilution factors and concentration factors, and provides examples and solutions for various dilution and concentration scenarios. It also covers the enumeration of microorganisms using the colony forming units (CFU) method.
What you will learn
Typology: Exams
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In most environments, the microbial density is usually too high or too low to obtain good results in the enumeration of microorganisms by a direct culture of the sample. This situation requires the dilution or concentration of the sample prior to carry out any study. Additionally, solid samples must be diluted for an easier handling, in this way; they can be treated as liquid samples.
In most cases, we work with decimal dilutions. The simplest case is the preparation of 10 ml of the 1:10 dilution of the sample. For this, 1 ml of sample is added to 9 ml of diluent; consequently, in 10 ml of this 1:10 dilution, 1 ml corresponds to the sample. Expressing by an equation:
1 ml of sample 1:10 dilution = 10 ml of 1:10 dilution (or 10-1^ dilution) 1 ml of sample + 9 ml of diluent
If a higher dilution of the sample is required, successive dilutions can be prepared. For example, if the 1:100 dilution is needed (or the 10-2^ dilution), it can be prepared adding 1 ml of 1:10 dilution to 9 ml of diluent, according to the following equation:
1 ml of 10-1^ dilution 1:100 dilution = 10 ml of 1:100 dilution (10-2) 1 ml of 10-1^ dilution + 9 ml of diluent
Or directly, adding 1 ml of sample to 99 ml of diluent:
1 ml of sample 1:100 dilution = 100 ml of 1:100 dilution (10-2) 1 ml of sample + 99 ml of diluent
Or adding 0.1 ml of sample to 9.9 ml of diluent:
0.1 ml of sample 1:100 dilution = 10 ml of 1:100 dilution (10-2) 0.1 ml of sample + 9.9 ml of diluent
Note: The final volume obtained is given by the denominator of the equation.
A colonies CFU/ml = X Dilution Factor B volume plated (ml) A colonies 1 CFU/ml = X B volume plated (ml) Concentration Factor
1.6. We have prepared the followings dilutions from different sample: 1:10, 1:5, 1:4 and 1:2. Then, 0.1 ml of each dilution is plated. After incubation, in all cases 27 colonies grew in the plates, which are the microbial densities of the samples?
1.7. A food sample was processed following this protocol:
SOLUTIONS
SAMPLE
1.1. 250 ml of a 10-^1 dilution
1.2. 10 ml of a 10-5^ dilution in 3 steps
There are other options, for example:
1 ml 0,1 ml 0,1 ml
9 ml 9,9 ml 9,9 ml 10 -1^10 -3^10 - 1:10 1:10 x 1:100 1:1,000 x 1: 1:1,000 1:100,
0,1 ml 0,1 ml 1 ml
9 .9 ml 9.9 ml 9 ml 10 -2^10 -4^10 - 1:100 1:100 x 1:100 1:10,000 x 1: 1:10,000 1:100,
