Measure Absorbance of M. xanthus Culture

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1. Turn on the spectrophotometer so it can warm up.

2. Obtain one 13mm glass test tube for each sample that you want to measure density for, plus one to blank.

3. Add 3mL of uncultured CTTYE growth media to the tube that will serve as your blank.

4. Add 2.5mL of uncultured CTTYE to labeled sample test tubes, and add 0.5mL of bacterial culture to the correspondingly labeled tube. **Note, if your sample is particularly dilute, add 1mL culture and 2mL nutrient media for dilution**

5. Blank the spectrophotometer with your blank tube.

6. One at a time, vortex one tube of diluted culture, then immediately record the Absorbance value at 600nm. Repeat for each sample.

7. Turn off the spectrophotometer if you are done and if no one else plans to use it relatively soon.

8. To obtain OD600 from these Absorbance values, divide by a path length of 1.3cm (diameter of the test tubes). Make sure you adjust this value for whatever dilution you performed in step 4.

9. You can use the below equation to calculate cells/mL of your sample:

Density (cells/mL) = 1.1x10^9 * (OD600) - 2.1x10^7

Note: for phenotype assays, use this cells/mL calculation to decide how to dilute or concentrate your sample accordingly.