Figure 2.
Example of a ddPCR-based strategy for detecting 2G TKI-resistant mutations at the time of switch. Representative 2-dimensional plots (in which channel 1 fluorescence [FAM] is plotted against channel 2 fluorescence [HEX]) expected for mutations conferring resistance to one or more 2G TKI. Black clusters at the bottom left corner represent FAM/HEX double-negative droplets. Green clusters at the bottom right corner correspond to droplets positive for e13a2 or e14a2 BCR::ABL1 fusion transcripts (detected by HEX-conjugated probes). Blue clusters at the top left corner correspond to droplets positive for the indicated BCR::ABL1 KD mutations or mutation subgroups (detected by FAM or FAM/HEX-conjugated probes). Orange clusters at the top right corner correspond to double-positive droplets. One tube is specific for the pan-resistant T315I mutation; one tube detects dasatinib-resistant mutations (with the nucleotide substitution leading to the V299L mutation clustering separately from the others since they also confer resistance to bosutinib); and one tube detects nilotinib-resistant mutations (with the E255K mutation clustering separately from the others since it also confers resistance to bosutinib). Results are expressed as percentage of mutant transcripts over total BCR::ABL1 transcripts. 2G TKI, second-generation tyrosine kinase inhibitor.

Example of a ddPCR-based strategy for detecting 2G TKI-resistant mutations at the time of switch. Representative 2-dimensional plots (in which channel 1 fluorescence [FAM] is plotted against channel 2 fluorescence [HEX]) expected for mutations conferring resistance to one or more 2G TKI. Black clusters at the bottom left corner represent FAM/HEX double-negative droplets. Green clusters at the bottom right corner correspond to droplets positive for e13a2 or e14a2 BCR::ABL1 fusion transcripts (detected by HEX-conjugated probes). Blue clusters at the top left corner correspond to droplets positive for the indicated BCR::ABL1 KD mutations or mutation subgroups (detected by FAM or FAM/HEX-conjugated probes). Orange clusters at the top right corner correspond to double-positive droplets. One tube is specific for the pan-resistant T315I mutation; one tube detects dasatinib-resistant mutations (with the nucleotide substitution leading to the V299L mutation clustering separately from the others since they also confer resistance to bosutinib); and one tube detects nilotinib-resistant mutations (with the E255K mutation clustering separately from the others since it also confers resistance to bosutinib). Results are expressed as percentage of mutant transcripts over total BCR::ABL1 transcripts. 2G TKI, second-generation tyrosine kinase inhibitor.

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