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MYD88L265P mutation detection by ddPCR on BM, PB and ctDNA
mutation quantification. Of note, 2 samples scored WT in all three compartments (Figure 4 and Online Supplementary Figure S6), while in 3 other patients MYD88L265P was detected in BM and PB, but scored WT in plasma ctDNA; these samples were excluded from the analysis since neg- ativity, confirmed by RNAseP analysis, was ascribed to
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Figure 2. Digital droplet polymerase chain reaction sensitivity test and exam- ple plots of controls (MUT vs. WT). (A) ddPCR assay measured on a 10-fold dilu- tion standard curve, shows a sensitivity of 0.0035% MUT (1 mutated copy in 30000 WT). (B) Example of ddPCR result plot for the MYD88L265P control. (C) Example of ddPCR result plot for the MYD88WT control. MUT: mutated; WT: wild- type.
pre-analytical issues, related to sample collection and stor- age. Two cases were erroneously collected in lithium- heparin tubes instead of K3EDTA, while one case was col- lected correctly but processed more than 6 h after the blood had been drawn, confirming the importance of col- lection tubes and processing timing for good quality ctDNA, as already reported.33–35
Minimal residual disease monitoring in Waldenström macroglobulinemia by MYD88L265P droplet digital polymerase chain reaction
To explore the potential role of MYD88L265P mutation detection by ddPCR for MRD monitoring and therapy response evaluation, we focused our analysis on 52 patients who had at least one available follow-up sample. These patients’ features and the treatment they received
Figure 3. MYD88L265P assay and IGH-based approaches are superimposable. Comparison of MYD88 and IGH-based minimal residual disease monitoring (expressed as copies of mutation in 1x105 cells) in 23 samples from 10 patients shows a good degree of correlation (r2=0.64).
are pre- sented in Online
Figure 4. Cell tumor DNA from plasma mirrors the bone marrow mutation level. MYD88L265P mutated/wild-type (MUT/WT) ratio in bone marrow (BM), peripheral blood (PB) and plasma (PL) paired samples from 32 patients show no statistical differences between BM and PL. The dashed red line shows the cut-off for mutational status. Symbols below the red line represent MYD88WT samples. RE: relapsed; NT: Naïve-to-Treatment patients.
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