You are given a culture Tube A of unknown concentration You

You are given a culture (Tube A) of unknown concentration. You make the following sequential dilutions: a. Transfer 2 ml of culture from Tube A to 38 ml of diluent in beaker B. Mix well. b. Transfer 1 ml of culture from beaker B to 4 ml of diluent in Tube C. Mix well. c. Transfer 100 l of culture from Tube C to 1.9 ml of diluent in Tube D. Mix well. d. Plate 400 l of the culture in Tube D onto a plate and incubate. The next day you fine 137 colonies on the plate. Draw out this dilution series and indicate the cumulative dilution factor at each step. Calculate the concentration in the original culture (Tube A). If you plate 100 l of the culture in Tube C, how many colonies should you expect? Be sure to show your work. Use diagram where they would be helpful.

Solution

a. Transfer of 2 ml from Tube A to 38 ml of beaker B

Vol in Tube A=2 ml, Vol in Tube B= 38+2= 40 ml, Dilution factor in B= 40/2=20

b. Transfer 1 ml from Tube B to 4 ml of Tube C

Vol in Tube B= 1 ml, Vol in Tube C= 4+1= 5 ml, Dilution factor in C= 5/1=5 and CDF = 20*5=100

c. Transfer 100 ml from Tube C to 1.9 ml in Tube D

Vol in Tube C= 100 ml, Vol in Tube D= 1.9 ml+100 ml=101.9 ml. DF in Tube D= 101.9/100= 1.019 and

CDF= 100 *1.019= 101.9

d. Placed volume=400 ml, Number of colonies formed= 137

Concentration of D tube = 137/400 = 0.34 cfu/ml

e. Concentration in Tube A (original)= 0.34 *101.9= 34.90 cfu/ml

f. 400 ml can give 137 colonies, so 1 ml will give 137/400 that is 0.34 colonies per ml

If there is 100 ml in tube C, it will give =0.34 *1/4*1.019 where 0.34 is colonies per ml, 1/4 because 100 is one fourth times of 400 , and 1.019 is DF between tubes C and D.

Answer is 0.0866