Hi. I'm currently working on my final year project, which involves microbiological and protein works.
I got a little confusion on the method to measure OD for my bacterial culture in order to construct a microbial growth curve. I got 2 methods here:
Method #1: Take a sample of 1 mL from the suspension culture and transfer into a cuvette. Then, measure the OD600 against the sterile broth as the blank.
Method #2: Take a sample of 1 mL from the suspension culture and transfer into a 1.5 mL centrifuge tube. Centrifuge the sample and remove the supernatant. Then, wash and resuspend the pellet with 1 mL saline and transfer into a cuvette. Measure the OD600 against distilled water as the blank.
Can someone explain to me regarding this matter? Thank you.
Khairil Ikhwan Ahmad Zahli, you could use either of the methods. The second method (Method #2) however ensures that you do not have interferences from other components of culture media (broth). Nonetheless, since in Method #1 you use the broth as your blank it could eliminate this worry. However, the broth used in this case is "fresh" and certainly would differ from that which the cells are suspended in during the measurement. So generally, I prefer Method #2 (though the result might not vary significantly when compared to Method #1).
Both method is correct, you can try any of them, I think method 1 is easy than 2, in method 1 you should not think about other components because you are using the sterile media for blanking that nullify other components OD. So generally, I prefer method 1.
Hi@ Both the methods are usually carried out in laboratories. Taking OD at 600nm implies that you are taking absorbance of live bacterial cells. When it is centrifuged chances of getting live bacterial cells may be less.Hence apply two methods, repeat thrice and compare. In addition go for subculture method using selective media for observation of CFU/ml
Good morning, I think both methods are fine, but I recommend cuantified the cells to the dilution method and plate pouring to have only living cells (CFU / mL).
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