Table 3.
Three-Day 5-Plate Dimethyl Sulfoxide Validation of the IL-6–Induced pSTAT3-Y705 Activation High-Content Screening Assay
| Plate controls average inner intensity pSTAT3-Y705 Ch 2 | Plate performance statistics | |||
|---|---|---|---|---|
| Plate ID | Mean_Max (n=32) | Mean_Min (n=32) | Z′-factor | S:B ratio |
| Day 1 | ||||
| 1 | 8,526.2 | 1,111.5 | 0.53 | 7.67 |
| 2 | 8,444.2 | 1,113.7 | 0.41 | 7.58 |
| 3 | 6,887.6 | 1,104.6 | 0.21 | 6.24 |
| 4 | 8,444.2 | 1,188.2 | 0.53 | 7.11 |
| 5 | 7,282.1 | 1,169.3 | 0.29 | 6.23 |
| Day 2 | ||||
| 1 | 7,797.2 | 1,519.8 | 0.62 | 5.13 |
| 2 | 7,767.1 | 1,372.3 | 0.59 | 5.66 |
| 3 | 7,843.9 | 1,378.5 | 0.69 | 5.69 |
| 4 | 8,401.7 | 1,481.0 | 0.62 | 5.67 |
| 5 | 9,203.9 | 1,558.2 | 0.62 | 5.91 |
| Day 3 | ||||
| 1 | 8,466.3 | 1,388.0 | 0.64 | 6.10 |
| 2 | 7,889.1 | 1,368.2 | 0.53 | 5.77 |
| 3 | 7,893.8 | 1,381.0 | 0.57 | 5.72 |
| 4 | 8,020.5 | 1,351.9 | 0.55 | 5.93 |
| 5 | 9,018.4 | 1,553.3 | 0.67 | 5.81 |
The average inner intensity pSTAT3-Y705 Ch 2 data presented in the table were generated in three independent experiments of 5×384-well DMSO validation plates. These experiments were designed to mimic 3 days of automated screening operations in the pSTAT3-Y705 activation HCS assay, save that DMSO was added to wells instead of compounds. The average inner intensity pSTAT3-Y705 Ch 2 data of the maximum (50 ng/mL IL-6 and 0.2% DMSO, n=32) and minimum (0.2% DMSO, n=32) plate controls from each of the 15×384-well plates are presented.