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Help with calculating CFU prior to infecting mice

CFU calculations dilution

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#1 kozl

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Posted 11 May 2018 - 01:31 AM

Hi,

My bacteria stock is at a concentration of 1x10CFU/ml (it's in 1ml total volume). I would like to infect mice with different CFU concentrations: 5x107 5x105x105x10each in a 40ul volume (in PBS). Could you please help me with the calculations? 

 

 

1. Is this correct? 

1x10 to 5x107    -> dilution factor is 2, 40ul/2= 20ul, so if I take 20ul from the 1x10CFU/ml stock into 20ul of PBS I will get 5x107  

 

2. When I take volume from my original stock =  1x10CFU/ml, in order to get the new concentrations, am I changing the original 1x10CFU/ml concentration? 

 

Thanks!


Edited by kozl, 11 May 2018 - 01:33 AM.


#2 bob1

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Posted 11 May 2018 - 06:14 AM

1) If you want those as a concentration (e.g 5 x107 CFU/ml), as you say in your post then use C1V1 = C2V2

 

C1 = 1x108, V1=?, C2=5x107, V2= 0.04 ml

 

V1= (5x10x 0.04) / 1x108

V1=...

 

If you want the amount (i.e you want to take 5x107 PFU total) to take from your stock (which is what your calculation suggests) then you use V=N/C

 

V= 5x10/ 1x108

V=... Note the answer is in ml. 

 

Note that if you want these as amounts in 40 ul, you can not do it - the concentration of your stock is too low. You can only do this if you want your working solutions to be concentrations.

 

2) The concentration of your stock doesn't change, just the amount of PFU left. For example, if you took out 500 ul from your ml, you will have taken out 5x107 PFU in total. However the remaining stock is still  at 108 PFU/ml, so you could take out. This is how concentrations work - they stay the same. This is why you can make a 1 M tris stock in 100 ml using 12.14 g of tris or use 121.14 g of tris for a 1 L stock - but the concentration of the final solutions is the same.

 

Make sure you do not confuse concentration and amount!



#3 kozl

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Posted 11 May 2018 - 10:43 AM

Thank you very much for your detailed explanation! 

I have more than 1 tube of the 1x10CFU/ml (1 mL per tube) and I am intrested in the second option that you mentioned (the amount = CFU). 

So if I understand correctly, for the 5x107 CFU I will take (5x107/1x108=0.5=) 500ul from the 1x10tube. I can then centrifuge this and reconstitute it in 40ul PBS and I will be able to infect the mice with this 40ul volume? 

 

Thank you!



#4 bob1

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Posted 11 May 2018 - 11:08 AM

Possibly, you will lose some titre with the pelleting and resuspension - I would also expect that amount of CFU to take up a significant proportion (i not all) of the 40 ul



#5 kozl

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Posted 12 May 2018 - 05:00 AM

Thanks!! I will test it out. 







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