Question

You are a microbiology lab technician and you need to quantify the viable cell concentration in your stock culture before beginning an experiment. Since you do not know the range, you perform a serial dilution over several orders of magnitude. You use the pour plate method and plate 1 ml for each dilution level, in duplicate. Give the most accurate value for the number of viable cells in the original stock culture, in CFU / ml. Just enter a number, do not type the units!

Since you want the practice, you attempt a count for all plates, even though some are likely too numerous to count. Only use values that give you the best accuracy. That means only use values that are a "countable number" of colonies (see text section).

Dilution series colony counts:

1:10: 860, 745

1:100: 123, 114

1:1000: 11, 15

1:10,000: 2, 4

Answer 1

Ans.

#1. Average original CFU in 1:10 dilution =

Average of [Observed CFU per mL inoculum / (dilution factor)] in two plates

                        = [{860 cell mL^-1 / (1/10)} + {745 cell mL^-1 / (1/10)} ] / 2

                        = 8025 cells/ mL

#2. Average original CFU in 1:100 dilution =

= [{123 cell mL^-1 / (1/100)} + {114 cell mL^-1 / (1/100)} ] / 2

= 11850 cells/ mL

#3. Average original CFU in 1:1000 dilution =

= [{11 cell mL^-1 / (1/1000)} + {15 cell mL^-1 / (1/1000)} ] / 2

= 13000 cells/ mL

#4. Average original CFU in 1:10000 dilution =

= [{2 cell mL^-1 / (1/10000)} + {4 cell mL^-1 / (1/10000)} ] / 2

= 30000 cells/ mL

# A colony count of 100-300 colonies per plate is generally accepted for CFU enumeration. The lower observed CFU count per mL than the actual CFU count in original sample is due to clumping of cells during serial dilution. However, there is no means by which the observed CFU can be greater than actual value.

So, the highest CFU count calculated in any one the plate is closed to the actual CFU count in original sample.

Therefore, CFU count in original inoculum sample = 30000 cells/ mL

= 3.0 x 10⁴ cells/ mL