Mouse lung CFU calculation

[ace]

We use one mouse lung lobe (out of five available) to homogenize in 1ml of saline and obtain CFU counts after infection with S.pneumoniae. After homogenization, 100ul of the homogenate is diluted in 900ul of saline (1:10 dilution) and plated. Now, I have been given to follow an established formula which says the dilution factor in this case is 400 (800 in case I homogenized in 2ml) - once the colonies grow, the counts are multiplied by this factor of 400 (4000, 40000....for subsequent dilutions) to obtain the total CFU. I am confused not sure how they arrived at this number of 400. Help!

[relaxin]

Me too.

You have just made one dilution (1:10), so the dilution factor should be 10. If you plate out only 100 ul (i.e. 1/10) of the diluted homogenate, you can calculate the total CFU for that lobe of lung tissue as "number of CFU on the plate X 100". But if you plate out only 25 ul (i.e. 1/40) of the diluted homogenate, then the total CFU will be "number of CFU on the plate X 400).
More accurate calculation should be expressed as "number of CFU/mg lung tissue". as the lobe comes in different size.

[ace]

Hi Relaxin,

Thanks for the reply. You are right about plating 100ul of the diluted homogenate. Apparently, the fifth lung lobe is not considered in these calculations. So, since you are using one lobe of the four (1/4th), the dilution factor of 100 is multiplied by 4= 400. The counts are usually expressed as log10 CFU/lung. Now is that a correct way doing the calculation?? I am more inclined towards your suggestion of doing it per mg of lung tissue. Any inputs are appreciated.

[relaxin]

I think either way will be OK, as long as you explain to the readers the basis of the dilution factor.

If I am the one who does the work, I would homogenize the whole lung in 4 ml saline, make dilutions and plate out aliquots in triplicates. Then the result can be truly expressed as CFU/lung. Multipling the result of one lobe by "4" to get CFU/lung is only an estimate.