Figure 2 -.

Similarity matrix generated in Morpheus. Columns were sorted by MOA, Compound, and then Concentration. (A) A subset of the similarity matrix showing the MOAs “Microtubule inhibitor” and “Microtubule stabilizing agent”. The top left and bottom right large red blocks show similarity of various doses on various plates within the same MOA class; the blocks on the top right and lower left are identical except for rotation, and show the similarity across classes. The small solid black box in the center shows the lowest dose of microtubule-stabilizing agent clusters well across replicates; its relatively poor correlation with the tightly clustered replicates at higher doses (black-dashed box) or any concentrations of microtubule inhibitor (green box) shows it might be below the effective dose of this drug. Higher doses of a microtubule-stabilizing agent, cluster well within and across doses, though a subtle recurring pattern within this block (highlighted by the yellow arrows) indicates that one of the five replicates shows a somewhat different profile than the other four, indicating a possible batch effect or technical anomaly. The effective concentration of a drug is highlighted by the lowest dose of ixabepilone clustering together (black box) but having weak correlations with the highest doses of ixabepilone. The higher doses of the microtubule-stabilizing agent are extremely similar to low concentrations of microtubule inhibitor (blue box) but less similar to higher concentrations of microtubule inhibitor (purple box). (B) Negative control (DMSO) correlation pattern, zoom out view of the similarity matrix. Black arrows highlight artifacts from plate-layout effects; treatments plated in the same or very similar well positions still can show significant similarity even after normalization. This can be alleviated at the experimental level by scrambling positions across plates and/or plating the same treatment in multiple positions spread across an individual plate.