How to Calculate CFU per ml of a Bacterial sample? In simple 3 steps

Correct Equation for Calculating

CFU/ml =  (No. of colonies x Total dilution factor)/Volume of culture plated in ml

CFU is the Colony Forming Unit

Here we discuss a very easy 3 step process for finding out CFU per ml of original stock?

Lets workout a question to understand the concept

This is the summarized video of this topic

Question: What is the number of CFU per ml of original stock? the following figure.

Step 1: Find out the plate with countable no. of colonies

Countable no. of colonies is a plate with colony count between 30-300. This number is experimentally and statistically taken as per standard lab manuals in microbiology.

In the Question given select a plate with number of colonies between 30-300.

In plate 1 colony count is 443; it is considered as “Too Many To Count” (TMTC); that is overcrowded therefore chance of error is high.

In plate 2 colony count is 63; between 30-300; this plate is to be taken for further calculation.

In plate 3 colony count is 3; it is considered as “Too Few To Count” (TFTC); that is small sample size; therefore chance of error is high.

So we have taken plate 2 with 63 numbers of colonies

Step 2: Find out the Total Dilution Factor (TDF)

Dilution Factor (DF) =Vf/Vi

Where Vf = Volume of diluent +Stock volume

            Vi=Volume of stock transferred

Lets do it for the first test tube a

DF=Vf/Vi=10/1=10 or10¹

For the first testtube DF and TDF is the same as we have diluted only once that is 10¹

For Test tube 2

DF=Vf/Vi=10/1=10 or10¹

Total Dilution Factor (TDF)=DF1xDF2XDF3

B tube=10¹x10¹=10²

C tube=10¹x10¹x10¹=10³

As a serial dilution procedure; it is very easy; ten times dilution at each step

So find out the TDF of the culture tube from which we cultured to get the plate with colony count 63

The TDF of plate with 63 colonies is 10⁵  

This is to be taken as TDF for further Calculation

Step 3: Substitute the Values in the Equation

No. of colonies=63

Total dilution factor=10⁵

*Vol. plated=0.1 ml

*Volume plated is 0.1ml that should be considered.

CFU/ml =  (No. of colonies x Total dilution factor)/Volume of culture plated in ml

              CFU/ml in original stock= 63x10⁵  /0.1 ml =63x10⁶  =63,000,000  CFU/ml

                                                                                              =6.3 × 10⁷ cfu/ml     

Here TDF is 10⁶ . From the culture tube with TDF 10⁵ , we have taken only 0.1 ml of 1 ml, that is further diluted to 10 times to become 10⁶ total dilution or TDF. 

Related Topic: Serial Dilution Protocol 

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