Materials and Reagents

1. Egg preparation
   100 eggs are used for each petri dish. Assays are performed in triplicate.
   
2. LB broth
   
3. Cholesterol
   
4. Ethanol
   
5. CaCl₂
   
6. MgSO₄
   
7. KPO₄
   
8. 3.7% (v/v) 1-octanol (Merck KGaA, catalog number: 820931 ) in Ethanol
   This concentration was established after a titration assay to find the lowest concentration needed for a fast response (within the minutes range) (Olmedo et al., 2012)
   
9. NGM plates (see Recipes)
   Note: For one experiment, always use plates that were poured from the same batch and were allowed to dry for the same amount of time.
   
10. Escherichia coli (E. coli) OP50 culture at 100 g wet bacterial weight/liter (see Recipes)

Equipment

1. Template for labeling plate (Figure 1)
   
   
   Figure 1. Template for plate labeling. On a piece of paper or plastic, draw a 5 cm-diameter circle in which to place the petri dish and a concentric 1 cm-diameter circle to mark the spot for the E. coli drop. Mark the center of this circle (A) with a dot. Draw another dot (B) at 0.8 cm from the center of the circle.
   
   
2. 5 cm Petri plates
   
3. Stereomicroscope equipped for picture acquisition (we use a Stereo Discovery V8 from Zeiss at a 10x magnification.)
   
4. Centrifuge
   

Procedure