It is very difficult to obtain a Colony Forming Unit by a single cell. I would like to know if it is possible to verify the presence of different cells from different bacteria.
Perform a preliminary count of each dish once the bacteria have had time to incubate, usually one or two days. You want to count only individual colonies, which should be distinct, isolated dots, not a whole blob of different colonies grown together. Choose the plate which has more than 30 of these colonies but less than 300.
Count the number of individual colonies. This is the CFU number of your dilution -- you will have to perform a simple calculation to determine the CFU of the original sample. For this example, a hypothetical plate containing 46 colonies will be used.
Determine the size of the dilution you used. Ideally, you labeled the petri dishes ahead of time. For this example, you can mix 1 mL of bacterial culture with 99 mL of saline. This is a 1/100 dilution.
Multiply the degree of the dilution by the amount you actually plated. If, in this example, you plated 0.1 mL of your 1/100 dilution onto the agar, you multiply 0.1 x 1/100, for a result of 1/1000 or 0.001.
Divide the CFU of the dilution (the number of colonies you counted) by the result from step 4. For this example, you would divide 46 by 1/1000, which is the same as multiplying by 1,000. The result is 46,000 CFU in the original sample.
References
- "Prescott's Microbiology"; Willey, Sherwood, Woolverton; McGraw-Hill 2010
The idea is this, if you have more than one type of cells in stock for examples; you have to make serial of dilution of that stock. Prepare proper growth medium and plate 0.1ml of that stock over the medium in petri dishe. Each cell with grow and form colony, the number of colony is the CFU. each colony has different appearance so you can differentiate , or do the same steps but use specific medium for each group
there are several methods (microfluidics, flow-cytometry etc.) which you can use to verify the number of cells in CFU. But they are comparatively complicated to the agar-plate method. Practically, it is the easiest and cheapest method.
Perform a preliminary count of each dish once the bacteria have had time to incubate, usually one or two days. You want to count only individual colonies, which should be distinct, isolated dots, not a whole blob of different colonies grown together. Choose the plate which has more than 30 of these colonies but less than 300.
Count the number of individual colonies. This is the CFU number of your dilution -- you will have to perform a simple calculation to determine the CFU of the original sample. For this example, a hypothetical plate containing 46 colonies will be used.
Determine the size of the dilution you used. Ideally, you labeled the petri dishes ahead of time. For this example, you can mix 1 mL of bacterial culture with 99 mL of saline. This is a 1/100 dilution.
Multiply the degree of the dilution by the amount you actually plated. If, in this example, you plated 0.1 mL of your 1/100 dilution onto the agar, you multiply 0.1 x 1/100, for a result of 1/1000 or 0.001.
Divide the CFU of the dilution (the number of colonies you counted) by the result from step 4. For this example, you would divide 46 by 1/1000, which is the same as multiplying by 1,000. The result is 46,000 CFU in the original sample.
References
- "Prescott's Microbiology"; Willey, Sherwood, Woolverton; McGraw-Hill 2010
Dear Istituto Superiore di Sanità:-
Colony counter is a suitable technique for counting the number of colony forming unit (CFU). Colonies more than 300 CFU and less than 30 CFU will be neglected as plates and re-working dilution and plating is recommended. if the colonies on the plates between 30-300 colonies, colony counter will be used as a simple method for detecting the number of colonies.
Other method is total viable count method by making a double dilution technique is also recommended to check the number of bacteria especially viable state bacteria.....Kind regard
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