Question

How do I calculate the dilution factor?

If you diluted your original cell culture by mixing 50 µL of your original culture in 450 µL of culture media, and then took 100 µL of that dilution and mixed it with 100 µL of Trypan blue, what is the dilution factor of your original culture?
I know dilution factor is DF = Vf/Vi
so my Vf is 50 +450 = 500
and Vi will be 50 giving me a dilution factor of 10.
but where does Trypan blue come from?

Answer 1

DF =`20`

Explanation:

The Trypan blue is just another reagent in the reaction. However, the important part for the dilution is that it also adds VOLUME to the solution, increasing the dilution.

The tricky part is that only a portion of the first dilution is taken, so the "initial" amount is reduced in the second dilution.

The only two critical values are the original volumes and the final TOTAL volume of the solutions.
FIRST dilution
`V_f = 50 + 450` ; `V_i = 50` Dilution Factor = `10`

SECOND Dilution
`V_f = 100 + 100` ; `V_i = 100` Dilution Factor = `2`

COMBINED Dilution Factor = `10xx2 = 20`

Note that if you had NOT taken a sample of the first dilution, the final volume would have been 600, for a dilution factor of
`600/50 = 12`