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**Figure 1. Isocitrate-dependent NADPH production and alpha-ketoglutarate-dependent NADPH consumption of wild-type or R172 mutant IDH2** IDH oxidative and reductive activity assays were performed on lysates of HEK293T cells transfected with wild-type or R172K mutant IDH2, or empty vector. **(A)** Lysates were assessed for generation of NADPH from NADP+ in the presence of 0.4 mM isocitrate over the indicated time course. Mean change in OD340 from the beginning of each assay (time = 0) is reported for each biological repeat performed [n=7] and error bars represent s.e.m. **(B)** Linear regression slopes were determined for each biological repeat of the isocitrate-dependent NADPH production assay. Box and whisker plot with median represented as the line through the box and whiskers representing values within 1.5 IQR of the first and third quartile. Means as black dot and bold error bars represent 95% CI. Linear regression slope determined from the data estimated from the representative experiment reported in Figure 2A of Ward et al., 2010 is displayed as a single point (red circle) for comparison. Statistical analysis was performed on data generated during this replication attempt. Wild-type IDH2 compared to vector control: Wilcoxon-Mann-Whitney test; *Z* = 3.13, uncorrected *p* = .00058, Bonferroni corrected *p* = .0023. R172K mutant IDH2 compared to vector control: Wilcoxon-Mann-Whitney test; *Z* = 1.60, uncorrected *p* = .13, Bonferroni corrected *p* = .51. **(C)** Lysates were assessed for consumption of NADPH in the presence of 1 mM alpha-ketoglutarate over the indicated time course. Mean change in OD340 from the beginning of each assay (time = 0) is reported for each biological repeat performed [n=7] and error bars represent s.e.m. **(D)** Linear regression slopes were determined for each biological repeat of the alpha-ketoglutarate-dependent NADPH consumption assay. Box and whisker plot with median represented as the line through the box and whiskers representing values within 1.5 IQR of the first and third quartile. Means as black dot and bold error bars represent 95% CI. Linear regression slope determined from the data estimated from the representative experiment reported in Figure 2B of Ward et al., 2010 is displayed as a single point (red circle) for comparison. Statistical analysis was performed on data generated during this replication attempt. Wild-type IDH2 compared to vector control: Welch’s t-test; *t*(9.42) = 0.29, uncorrected *p* = .78, Bonferroni corrected *p* > .99. R172K mutant IDH2 compared to vector control: Welch’s t-test; *t*(6.95) = 5.97, uncorrected *p* = .00058, Bonferroni corrected *p* = .0023. **(E)** **Representative Western blots probed with an anti-IDH1 antibody, an anti-IDH2 antibody, and an anti-alpha-Tubulin antibody. Additional details for this experiment can be found at [https://osf.io/6ve4d/](1). ** **(E) Representative Western Blot images are as follows and can be found along with all western blots for this protocol [here](2):** - 160725 IDH2 assays IDH2 3min2.jpg (IDH2) - 160725 IDH2 assays IDH1 5min'.jpg (IDH1) - Tubulin 2min JASON.jpg (TUB) Access [full size image][1] and [R script][2] used to generate this figure. This script will call the [data][3] directly from the OSF. <br> <br> ---------- [Statistical Analyses component][4] contains R scripts that were used for the analysis. [1]: https://osf.io/pdsr9/ [2]: https://osf.io/t5gf9/files/ [3]: https://osf.io/xxjje/ [4]: https://osf.io/vupmw/wiki/home/