TABLE 2.
Comparison of assay conditions of skin sensitization in vitro assays and their qHTS versions.
| Culture time | Exposure time | Maximal test concentration | Negative/positive threshold | |
|---|---|---|---|---|
| KS | 24 h | 48 h | 5,000 mg/L | 1.5-fold induction |
| qHTS KS | 5 h | 24 h | 92 µM | Curve rank number >3 in curve rank criteria |
| 1.5-fold induction in OECD TG criteria |
| C/K-peptide concentration | Exposure time | Test method | Prediction model | |
| DPRA-Cys | 0.667 mM in pH 7.5 phosphate buffer | 24 h | HPLC | Negative: Cys % depletion <13.89% |
| Low reactivity: 13.89 < Cys % depletion <23.09% | ||||
| Moderate reactivity: 23.09 < Cys % depletion <98.24% | ||||
| High reactivity: Cys % depletion >98.24% | ||||
| SPE–MS/MS DPRA-C | 0.5 µM in pH 7.5 phosphate buffer | 24 h | MS-MS | Same as above |
| DPRA-Lys | 0.5 mM in pH 10.2 ammonium acetate buffer | 24 h | HPLC | Negative: mean Cys and Lys % depletion <6.38% |
| Low reactivity: 6.38% < mean Cys and Lys % depletion <22.62% | ||||
| Moderate reactivity: 22.62% < mean Cys and Lys % depletion <42.47% | ||||
| High reactivity: 42.47% < mean % depletion <100% | ||||
| SPE-MS/MS-DPRA-K | 0.5 mM in pH 10.2 ammonium acetate buffer | 24 h | MS-MS | Same as above |
| Culture time | Exposure time | Test concentration | Negative/positive threshold | |
| hCLAT-CD86 | 24 h | 24 h | Up to 5,000 mg/L | RFI 150 |
| CD86 | 24 h | 24 h | Up to 100 µM | RFI 150 |
| hCLAT-CD54 | 24 h | 24 h | Up to 5,000 mg/L | RFI 200 |
| CD54 | 24 h | 24 h | Up to 100 µM | RFI 200 |
| Cell line | Exposure time and test concentration | Assay type | Negative/positive threshold | |
| IL8-Luc | THP1-Luc | 24 h, 20 mg/mL | Luciferase reporter assay | 1.5-fold |
| HTRF IL8 | THP1 | 16 h, up to 100 µM | HTRF immune assay | 1.5-fold |