Table 2.
Protocol summary and assay statistics
| Parameter | Description or value |
|---|---|
| Protocol summary | |
| Growth medium and microtiter plates | Clear-bottom plates (384 wells) containing 50 μl of LB medium with 0.3% agar |
| Compound usage | Compounds were dispensed to the top of the soft agar and allowed to diffuse for 2–4 h at room temp |
| Inoculation | A 2.5-nl sample containing ∼20 CFU of strain C7258 was dispensed into the bottom left corner of each well (see Fig. S3 in the supplemental material) |
| Incubation | Plates were incubated for 16–24 h at 30°C and high humidity |
| Motility reading | Spreading of motile bacteria was measured by reading the OD615 above and to the right diagonal of the inoculation site (see Fig. S3) |
| Viability reading | Viability was estimated by adding 5 μl of 100% alamarBlue to each well; plates were incubated for 1–1.5 h at 30°C, and fluorescence was read (excitation wavelength, 535 nm; emission wavelength, 595 nm) |
| Control drugs | Phenamil (motility) and tetracycline (viability) |
| Assay statistics | |
| Z value (mean) | 0.5–0.8 (0.7) |
| Signal-to-noise ratio | 173 |
| Signal-to-background ratio | 7 |
| Pilot screen | |
| No. of compounds screened | 8,093 (in duplicate on two different days) |
| Control OD615 (avg [coefficient of variation]) | |
| Carrier control (DMSO) | 0.73 (5.6) |
| Motility control (phenamil) | 0.10 (3.2) |
| Viability control (tetracycline) | 0.10 (3.6) |
| Hit rate (%) | 0.2 |