8: Why we plate more than one dilution?

500 in one dish

Remember the picture of the impossible-to-count Petri dish with 500 colonies? No one wants that to happen to them, especially late on a Friday afternoon. So, how can we avoid it?

Very easily … while we’re doing our dilutions, we just keep plating each intermediate step. Then the next day, we decide which plate looks the most reasonable for counting. Since we cleverly labeled each plate with its overall dilution, we also know how to scale back up to get the original concentration.

So, now we finally have all the pieces of serial dilution assembled:

  • Dilute
  • Calculate dilution
  • Plate
  • Count colonies

To make things easier, the standard operating procedure is to go by factors of 10, and to do about 5 or 6 plates altogether.

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Let’s say I guesstimate that I have about 42 million CFUs per mL — I might want to start with a first dilution of about 1/1000 (or 10-3), and plate that. Then I make 4 more 1/10 (or 10-1) dilutions, plating each one in turn. In the end, I have 5 plates, labeled as shown below. And, if you roll over the image, you will see the plate counts I expect if my original guess (42 million) was approximately correct. Oh, and TMTC means “Too Many To Count”.