Extended Data Fig. 2: Practical solutions to minimize bias introduction.

(a) Schematic overview of all strategies deployed. (b) PathoPlex is compatible with multiple microscopy systems, including widefield, spinning disk and laser confocal, which directly determine imaging time, file size and image resolution. (c) Strategies to minimize bias introduction are based on creating self-contained batches, where tissues are processed in parallel using imaging chambers that contain specimens representing all experimental groups, including controls. (d) Suggested imaging chambers for histopathological studies (2–24 wells). (e) Multi-well pipetting can be performed manually or using liquid handling systems. Alternatively, 3D printing can be used to customize a 1-well imaging chamber, which simplifies liquid handling. (f) In addition, a 3D printer can also be used as a liquid handling system. The printer head is used to automate liquid addition and removal. (g) Specificity, and elution controls of 3D printer-based imaging cycles. LMNB1, Lamin B1 (cycle 1); EMCN, Endomucin (cycle 1); α-SMA, alpha-smooth muscle actin or ACTA2 (cycle 2); VMT, Vimentin (cycle 4); AKAP12, A-Kinase Anchoring Protein 12 (cycle 4); Secondary ab (QC cycle; cycle 6). Scale bars represent 100 µm. Parts of panels a, c and e were created using BioRender (https://biorender.com).