Table 1. A subset of assay parameters evaluated during validation.
| Parameters |
Result |
|
|---|---|---|
| AAV5 TAb assay | AAV5 TI assay | |
| Sensitivity (neat concentrations in plasma) | 4.9 ng/ml of a polyclonal anti-AAV5 (or 68.9 ng/ml ADK5a equivalents) | 65.9 ng/ml of a monoclonal anti-AAV5 (ADK5b) |
| Specificity | No cross-reactivity of 4 μg/ml anti-AAV8 | No cross-reactivity of 48 μg/ml anti-AAV8 |
| Cut point | 1.15 (SCP); 1.30 (TCP); 39.7% (CCP) | 44.9% Transduction (ACP) |
| Interassay precision (%CV) | HQC 14.3% | TQC 3.78% |
| LQC 13.1% | LQC 17.5% | |
| NQC 9.9% | NQC 35.6% | |
| Intra-assay precision (%CV) | HQC 4.6% | TQC 3.29% |
| LQC 7.2% | LQC 11.6% | |
| NQC 6.3% | NQC 13.2% | |
| Selectivity | 80% (8/10) pass at 0 ng/ml anti-AAV5 | 100% (10/10) pass at 0 ng/ml anti-AAV5 |
| 100% (10/10) pass at 9 ng/ml anti-AAV5 | 100% (10/10) pass at 180 ng/ml anti-AAV5 | |
| Interference, hemolysis | 100% (6/6) pass at 0 ng/ml anti-AAV5 | 66.7% (6/9) pass at 0 ng/ml anti-AAV5 |
| 100% (6/6) pass at 9 ng/ml anti-AAV5 | 100% (9/9) pass at 180 ng/ml anti-AAV5 | |
| Interference, lipemia | 66.7% (4/6) pass at 0 ng/ml anti-AAV5 | 33.3% (3/9) pass at 0 ng/ml anti-AAV5 |
| 100% (6/6) pass at 9 ng/ml anti-AAV5 | 100% (9/9) pass at 180 ng/ml anti-AAV5 | |
Abbreviations: %CV, percentage coefficient of variation; AAV, adeno-associated virus; ACP, assay cut point; CCP, confirmation or specificity cut point; HQC, high-quality control; LQC, low-quality control; NQC, negative quality control; SCP, screening cut point; TAb, total anti-capsid antibody; TCP, titer cut point; TI, transduction inhibition; TQC, titer quality control.