AML-87-001
We used a novel microfluidic approach with molecular barcode technology to amplify the DNA from individual cells. Briefly, cryopreserved bone marrow mononuclear cells were thawed, and cells were quantified using a Countess Automated Cell Counter (Thermo Fisher Scientific). The cells were resuspended in cell buffer and diluted to a concentration of 2,000,000 to 4,000,000 cells/mL. Next, 35-100 L of cell suspension was loaded onto a microfluidics cartridge and cells were encapsulated on the Tapestri instrument followed by the cell lysis and protease digestion on a thermal cycler within the individual droplet. The cell lysate was then barcoded such that each cell had a unique label. The barcoded samples were then thermocycled using either 50 primer pairs specific to a panel of 19 mutated genes covering known AML-related hotspot loci and 10 commonly heterozygous SNP loci for ADO determination (19-gene panel), or 279 primer pairs specific to a panel of 37 mutated cancer genes (custom-designed panel).