Genomic DNA was extracted from cell lines and human PB samples using standard methods. DNA was amplified (25 ng template per well) using different qPCR assays designed to detect wild-type JAK2 (JAK2-WT) and the JAK2 1849G>T mutation encoding JAK2-V617F (designated JAK2-V617F assay), in parallel with independent control gene assays for Cyclophilin A38 (link) and Albumin (ALB),39 (link) providing a control for any variation in the amount of template DNA between reaction wells, as well as between successive quality control (QC) rounds (see Supplementary Table 1 for primer and probe sequences). The evaluation encompassed a number of unpublished ‘in-house' assays (n=3), together with a large range of published assays (n=6).26 (link), 40 (link), 41 (link), 42 (link), 43 (link), 44 (link) The latter included assays that have been widely applied in clinical practice, that is, those forming the basis of the MutaQuant kit (Qiagen, Marseille, France),41 (link) or reported to predict outcome and used to guide management following allogeneic transplantation.26 (link), 29 (link), 31 (link), 32 (link) For published assays, reported reaction conditions were used; whereas for ‘in-house' assays, the reaction conditions used were provided by the respective source laboratories. Assays were run in duplicate in the QC rounds and in triplicate wells for the analysis of control PB and primary patient samples with appropriate water controls.