Table 2.
Validation results of analysis of anti-CD3/anti-CD28 bead contamination.
| Validation part | Parameter | Acceptance | Result | Passed | |
|---|---|---|---|---|---|
| Reproducibility | Coefficient of variation | ≤25 | 750 1,000 3,000 |
7.1 8.7 22.7 |
X yes □ no |
| Intermediate precision | F-value | <9.28 | 750 1,000 3,000 |
1.54 5.50 2.49 |
X yes □ no |
| Deviation from mean | ≤15% | 750 1,000 3,000 |
6.5 14.1 7.3 |
X yes □ no | |
| Coefficient of variation | ≤25 | 750 1,000 3,000 |
14.0 13.3 17.6 |
X yes □ no | |
| Linearity | Coefficient of correlation | ≤0.98 | 0.996 | X yes □ no | |
| Range | Detection range | 300–6,000 | 300–6,000 | X yes □ no | |
| Accuracy | Mean | 637–863 850–1,150 2,400–3,600 |
750 1,000 3,000 |
689 1,105 3,420 |
X yes □ no |
| Retrieval rate | ±15% ±15% ±20% |
750 1,000 3,000 |
8.1% 10.5% 14.0% |
X yes □ no | |
| Limit of detection | QL | <400 | 390 | X yes □ no | |
| Specificity | Positive particle | >400 | 1,105 | X yes □ no | |
| Negative particle | <400 | 38 | X yes □ no | ||
A total of 750, 1,000, and 3,000 anti-CD3/anti-CD28 expander beads were mixed with 100 × 106 peripheral blood mononuclear cells. For reproducibility, triplicate analysis of indicated cell-bead mixtures was statistically analyzed by coefficient of variation. The analysis of a different person on a different day was used to determine the intermediate precision. To determine linearity, 10 different cell-bead mixtures were analyzed by correlation factor and linear regression. These results were compared with the theoretical real fraction contents to determine accuracy. The limits of detection were determined by analyzing six bead-negative cell mixtures and calculated with the formula: 10s/a (s indicating SD and a indicating slope of regression). Specificity was determined by using bead-positive and bead-negative cell mixtures, respectively.
QL, quantification limit.