**Day 0**

1. Overnight yeast to be imaged.

2. Make serial dilutions - add 3.3 mL to tube A, 3 mL to tube B, 2.7 to tube C. Add sample to tube A and vortex. Take 300 uL from tube A, add to tube B, vortex. Take 300 uL from tube B, add to tube C, vortex.


**Day 1**

1. Label flip top tubes for each of your samples

2. Put 1 mL of appropriate overnight into each of the tubes

3. Centrifuge cells at 3600 rpm from 2 minutes at room temperature. Remove the supernatant

4. Resuspend cells in 1 mL of appropriate media

5. Add 5 uL of 0.2 M PSMF where appropriate (based on that day's experiments). PMSF is in the -20°C freezer

6. Add any drugs or vehicles where appropriate

7. Transfer to new, labeled culture tubes and shake at 30°C for 4 hours. SET A TIMER