Differences between multicolor flow cytometry–, RQ-PCR–, and HTS-based detection methods
| . | Multicolor flow cytometry . | RQ-PCR . | HTS . |
|---|---|---|---|
| Sensitivity limit of detection | 4-color flow: confirmed 10−4 | Confirmed 10−5 | Reported 10−6 |
| 6-color flow: reported 10−5 | |||
| Method | Surface antigen detection by different antibody combinations (eg, CD5/CD19/ CD20/CD43/CD79b/CD81) | Detection of disease-specific IGHV using patient-specific primers | Detection of disease-specific IGH sequences after amplification of all IGH gene segments using consensus primers |
| Fresh material required? | Yes; samples must be <48 h old | No, but DNA extraction preferably <48 h | |
| Standardized protocol? | Yes | Yes | Ongoing |
| Advantages | Directly quantitative | High sensitivity | High sensitivity |
| Widely available | No live leukocytes required | No live leukocytes required | |
| Results quickly available | Multiple mutations can be detected in 1 test | ||
| Highly standardized assay | |||
| Disadvantages | 4-color flow: lower sensitivity | Not directly quantitative | Not directly quantitative |
| Samples must be fresh | Requires baseline sample | Requires baseline sample | |
| Time and labor intensive | Less widely available | ||
| Expensive |
| . | Multicolor flow cytometry . | RQ-PCR . | HTS . |
|---|---|---|---|
| Sensitivity limit of detection | 4-color flow: confirmed 10−4 | Confirmed 10−5 | Reported 10−6 |
| 6-color flow: reported 10−5 | |||
| Method | Surface antigen detection by different antibody combinations (eg, CD5/CD19/ CD20/CD43/CD79b/CD81) | Detection of disease-specific IGHV using patient-specific primers | Detection of disease-specific IGH sequences after amplification of all IGH gene segments using consensus primers |
| Fresh material required? | Yes; samples must be <48 h old | No, but DNA extraction preferably <48 h | |
| Standardized protocol? | Yes | Yes | Ongoing |
| Advantages | Directly quantitative | High sensitivity | High sensitivity |
| Widely available | No live leukocytes required | No live leukocytes required | |
| Results quickly available | Multiple mutations can be detected in 1 test | ||
| Highly standardized assay | |||
| Disadvantages | 4-color flow: lower sensitivity | Not directly quantitative | Not directly quantitative |
| Samples must be fresh | Requires baseline sample | Requires baseline sample | |
| Time and labor intensive | Less widely available | ||
| Expensive |