Fig. 2

Validation of detection of amplifications in smMIP-based NGS analysis on gDNA from clinical FFPE specimens. a Relative coverage and z-scores in 5 positive controls for EGFR, MET and ERBB2 (n = 3: high, medium, and low level amplification) normalized to a normal tissue control series. Values of all 13 genes relevant for amplification detection (see Supplementary Table 1) are plotted for the five samples. Values were calculated per gene per sample and sorted by increasing value. The positive control values (one gene per samples) are depicted in green, values for all other genes (12 per samples) are shown in black. The additional detected EGFR amplification, shown in orange, was confirmed by FISH. b Grouped relative coverage in a series of 46 clinical tissue samples and 15 normal tissue controls. Values were calculated per gene per sample. c Relative coverage per gene in the series of 46 clinical samples, with additional clinical/molecular information (details in main text). The cut-off for validation (relative coverage ≥3.0) is shown by an orange line. Potential amplifications in green were validated by OncoScan array analysis (the others were not analyzed by OncoScan array). d Three positive control samples were diluted in gDNA isolated from normal tissue. Relative coverage (y-axis) and z-scores (above bars) compared with a normal tissue control series are shown. On the x-axis the dilution based on gDNA concentration is shown. EGFR positive control: unknown tumor purity. ERBB2 high positive control: 70% tumor cells. ERBB2 low positive control: 50% tumor cells