Table 3.

Cell culture devices qualification report. After developing topographic cell culture devices based on the guideline, researchers should report the controlling of confounding factors.

1. Structures Fabrication Related Confounding Factors Testing ☐ Well-controlled “one structural parameter” difference Supporting testing results: e.g., SEM demonstrated that the shapes of nanopores are similar, pore size is the only one variable structural parameter (50 nm, 100 nm, 200 nm), and other structural parameters are consistent between different topographies (pore depth: 50 nm; pore interval: 100 nm). ☐ Minimum and consistent chemical properties Supporting testing results: e.g., XPS and FTIR spectra showed no unwanted chemical elements or bioactive functional groups, and similar peaks between different topographic structures; ☐ The consistent structure-independent difference in physical properties Supporting testing results: e.g., AFM show similar Young's modulus between different substrates

2. Cell Culture Related Confounding Factors Testing ☐ Geometrical and physicochemical stability during the sterilization process Supporting testing results: e.g., SEM showed no obvious change in topography after sterilization. XPS, FTIR, and AFM proved no physicochemical alteration after sterilization. ☐ Geometrical and physicochemical stability during immersion Supporting testing results: e.g., SEM showed no obvious change in topography after immersion. XPS, FTIR, and AFM proved no physicochemical alteration after immersion. ☐ Immobile during immersion Supporting testing results: e.g., It can be visible that the substrates wound not float during immersion ☐ No structure-independent cytotoxicity Supporting testing results: e.g., CCK-8 showed good cell viability in the soaking medium of substrates

3. Cell Response Analysis Related Confounding Factors Testing ☐ For light microscopy: Transparency Supporting testing results: e.g., It can be visible that the substrates were transparent ☐ For confocal microscopy: No fluorescent interference Supporting testing results: e.g., Confocal microscopy exhibited no fluorescent interference ☐ For SEM: Non-magnetic Supporting testing results: e.g. XPS showed no magnetic elements in the cell culture device ☐ For hydrodynamic flow assay: Immobile during shear stress Supporting testing results: e.g., It can be visible that the substrates wound not float under shear stress ☐ For wound-healing assay: Geometrical stable after wound preparation Supporting testing results: e.g., SEM showed no change of the topographies after wound preparation. ☐ For CCK-8, MTT assays: Not to bring absorbance difference Supporting testing results: e.g., CCK-8 showed no confounding absorbance ☐ For RT-qPCR, WB assays: Not affect sample quality Supporting testing results: e.g., Spectrophotometry demonstrated a good quality of the mRNA extracted from the devices (A260/A280 > 1.8, A260/A230 > 1.8). ☐ For RT-qPCR, WB assays: Not affect internal reference Supporting testing results: e.g., we chose GAPDH as an internal reference, which stably expressed on different topographic structures.