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. Author manuscript; available in PMC: 2023 Apr 14.
Published in final edited form as: Nat Protoc. 2023 Jan 6;18(4):1243–1259. doi: 10.1038/s41596-022-00792-6

Fig. 7.

Fig. 7.

Cytocompatibility study using surfaces that were 3D-printed using the PEG-DA-258 resin with 0.4% (w/w) photoinitiator (IRG). Prior to seeding, we exposed the surfaces to a UV bath in water for 12 h so as to leach out possible un-reacted PEG-DA monomers and/or photoinitiator. We then treated the surfaces with oxygen plasma right before coating proteins or seeding cells. (a) Transparent PEG-DA-258 Petri dishes (25 mm-diam., 5 mm-tall and 1 mm-thick walls). (b) Phase-contrast image of Chinese hamster ovary cells (CHO-K1) seeded in a 3D-printed Petri dish. CHO-K1 cells were cultured in DMEM media (Invitrogen) supplemented with 10% fetal bovine serum (Hyclone) and grown in a 5% CO2 atmosphere at 37 °C. This image demonstrates that our 3D-printing technique is adequate for phase-contrast microscopy, a mode of imaging that is very sensitive to surface irregularities. (c, d) Fluorescence images of the cells cultured in the 3D-printed PEG-DA-258 Petri dish after labelling with live fluorescent tracer Cell Tracker Green (c) and Orange (d), showing that our resin yields a plastic that is appropriate for fluorescence-based observations. (e) Phase-contrast image of CHO cells seeded in a tissue culture polystyrene (TCPS) Petri dish as a control, showing no appreciable morphological differences compared to (b). (f) Phase-contrast image of primary mouse hippocampal neurons (embryonic day 818) seeded in a 3D-printed Petri dish. These surfaces were coated with poly-D-lysine (100 μg/mL) (Sigma-Aldrich) overnight and Matrigel (BD Biosciences) (diluted 1:60 with DMEM) for 1 h at 37 °C. Primary neurons were harvested from the hippocampi of embryonic day 18 mice (Brainbits), and enzymatically dissociated using a papain dissociation kit (Worthington Biochemical), following well established protocols. The dissociated cells were suspended in Neurobasal media (Invitrogen) supplemented with 1× B-27 (Invitrogen), 0.5 mM GlutaMax (Invitrogen) and 100 U/mL penicillin–streptomycin (Invitrogen), and plated onto the Matrigel-coated PEG-DA-258 surfaces.