Christina is perfectly right. However, what about up-to-date methods? For my experiments I am using DAPI counting.

CFU make sense for medical research with some bacteria or for the analysis like an old fashion API 20, however, in many systems you are observing the top of the iceberg even in the culture. (My experience is with V. cholerae typical for viable-but-not-cultivable cells).

If you need the culture for experiments, your curve would be out of standard because of the uncertain state of the cells in the agar-plate colony. I recommend you to pick up the colony, cultivate it in a small volume (10mL) for several hours (depending on the species; 6 h at 37°C may be OK) and already exponentially growing culture of known volume inoculate to your standard vessel. Sure, you should measure OD frequently. In routine experiments it works excellent; using 10mL in 500mL overnight, the bacteria disappearance in experimental water (low in nutrients) follows the same curve.

Other thing: OD is valid only for a limited interval (if you are not using nephelometry) because of the nature of the sample (particles).

Good luck

Mirek Macek

It is often important to know not only what types of bacteria are in a sample but also how many of them are present.

Additional information available in:                             http://classes.midlandstech.edu/carterp/courses/bio225/chap06/Microbial%20Growth%20ss5.htm

http://classes.midlandstech.edu/carterp/courses/bio225/chap06/Microbial%20Growth%20ss5.htm

References

You can do a standard curve of bacterial growth in a comparative way using both OD and CFU/ mL.First , subculture the pure bacterial strain in BHI broth till get young exponential cells , From these  and at sterile conditions pipette 100 micro litre cell suspension to 1.9 mL sterile BHI broth and measure the OD and CFUl/mL and use it as an initial step. Repeat when you get on 0.1 OD and comparable CFU/mL and make another trials and every 2 hours measure both OD and analyse CFU/mL , Then draw the standard curve between time and OD one one side and time with CFU/mL from another side.  Good LUCK

Hello Sarah, 

You can also make the calibration curve from a culture of 24 h on agar, to make different suspensions in a range of D.O (0.08 - 1.5), each of the adjusted tubes make dilutions for triplicate to count the CFU/mL. The data can be grouped in Excel, and based on the linear equation Y = mx + b, you determine the slope and the intercept and the coefficient r^2. This will allow you get in the formula to have according to a D.O the CFU/mL. Good Luck! 

You need to go back a step and ask what you will be using the OD/cfu information for. If you need a certain cfu/ml for a future experiment then what growth phase do you want the cells to be in? Gamal's suggestion of performing a growth curve experiment is the best place to start. Be sure to use a volume of medium and culture flask that allows sufficient gas transfer e.g. no more than 50ml medium in a 250ml flask (or equivalent), less if possible. I used to inoculate to an OD600 of 0.1 from an overnight culture then sample depending on the expected growth rate (every hour is a good starting point). For each sample, measure OD and plate for cfu (including the inoculated flask at time zero). Plot LnOD v time, you should get the classic lag-exponential-stationary plot. You can then calibrate OD/cfu for different growth phases. Make sure you continue the growth curve well into stationary phase, you will likely see a stable OD but decreasing cfu as cells die. Hope this helps!

You can find in the following good answers:

http://www.microbiol.org/resources/monographswhite-papers/measurement-of-cell-concentration-in-suspension-by-optical-density/