Figure 3

FRESH bioprinting using the open source bioprinter. (a) Design of the construct to be printed using FRESH methodology, and (b) actual FRESH bioprinted construct. (c) 3D rendering of a Col-F stained bioprinted collagen construct based on imaging by confocal microscopy. (d) Example of MDA-MB-231 breast cancer cells seeded onto a bioprinted collagen construct post printing. The cells were stained with SiR-actin and NucBlue to visualize the actin cytoskeleton and cell nuclei respectively. The cells were shown to attach and adopt normal cell morphologies a few hours after cell seeding. The relative level of active apoptosis (e) as measured by Caspase 3/7 activity, and relative occurrence of dead cells (f) as measured by propidium iodide signal in cells grown in glass-bottomed wells or on FRESH bioprinted collagen constructs, in the presence or absence of the apoptosis-inducing toxin staurosporine (10 μM for 5 h) are shown. (g) Analysis of cell viability as assessed by fluorescein diacetate (live) and propidium iodide (dead) staining of MDA-MB-231 breast cancer cells after overnight culture (~ 20 h) on bioprinted collage constructs, and 1 week later. (h) Example of fluorescein diacetate (live cells) and propidium iodide (dead cells) staining of MDA-MB-231 breast cancer cells distributed on the bioprinted collagen constructs following 1 week of culture. Differential interference contrast imaging (DIC) reveals the crisscross patterning of the construct.