Endy:F2620/Transfer Curve/Protocols

1. Prepare 6 cultures using the standard chassis prepartion protocol.
2. The standard protocol was modified by pre-adding the appropriate concentration of Endy:F2620/AHL to the wells. Each of the 3 replicates of the 6 cultures were induced with 6 concentrations of AHL (0nM, 0.01nM, 0.1nM, 1nM, 10nM, 100nM, 1μ M, 10μ M).
3. These plates were incubated in a Wallac Victor3 multi-well fluorimeter at 37C and assayed with an automatically repeating protocol of absorbance measurements (600nm filter, 0.1 second absorbance through approximately 0.5cm of fluid), fluorescence readings (488nm excitation filter, 525nm emission filter, 0.5 second, CW lamp energy 12902 units), and shaking (1 mm, linear, normal speed). One well was filled with M9 media in all experiments as an absorbance background. Time between measurements was 3 minutes.