I was making competent cells the other day with a culture volume of 100 mL (diluted from an O/N) in a 500 mL flask.
Starting at 1 h, I started measuring OD to check when the cultures reached OD600 of 0.4. I use a BioWave CO8000 cell density meter. I take 500 uL of culture and measure it against a blank of the medium.
But in the time I take it to the bench and then remove 500 uL, is that measurement really going to be accurate? Is there a better way to monitor the OD of a large culture? Would I be able to set up a smaller culture in a 15 mL tube at a similar dilution such that I can just take the reading from the culture tube? Please advise. Thank you!
Hi there,
I don't really understand your point... If you want to monitor the growth by spectrophotometry you have to withdraw aliquots to make the readings whatever the total volume of the culture... You have to make sure the suspension is homogenous and that the reading is within the window of linearity of the spec.
By the way growth rate of cultures made in large flask and in a small tube will be different because conditions will be different (mainly due to aeration and shaking) even if the starting point is the same.
Dear Omar Ocegueda
i think that your approach is ok.
I will not be worried about the sample homogeneity since the culture is shaked but in case you are afraid that i single reading is not enough precise you can try to take for each time more than 1 point (e.g 3x500ul) and make the average of it.
i'm not agree with the idea of set up a smaller culture in a 15 mL tube at a similar dilution such that I can just take the reading from the culture tube, since the oxigenation of the 2 cultures could be different and therefore also the growth rate.
best
Manuele
If in doubt make serial dilutions and measure those. If you chill your culture in a fridge the OD should not change much in the time you take to measure the OD.
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