Fig. 1 The confiner coverslip setup. (A) Left, diagram of the confiner coverslip setup. A confiner coverslip (blue) with spacers is placed on top of adherent or suspended cells (yellow). Right, picture of a 12-mm diameter confiner coverslip. (B) Side view of the confiner coverslip setup. Top, unconfined position. Bottom, confined position. Cm, culture medium. (C) Side view of the single-well dynamic confiner setup. Top, unconfined position (low vacuum pressure). Bottom, confined position (high vacuum pressure). Mc, metallic connector. Vp, vacuum pump. Pd, Petri dish. Cc, confiner coverslip. Gc, glass coverslip. Cm, culture medium. (D) Side view of the multi-well static confiner setup. Top, unconfined position. Bottom, confined position. Pc, Petri dish cover. Pd, Petri dish. Cm, culture medium. Gc, glass coverslip. Cc, confiner coverslip. Cp, PDMS confiner piston. Sp, PDMS spacer. Tp, tape.
Fig. 2 Cell confinement combined with nano-grooved surfaces. (A) Side view of the confiner coverslip setup combined with nano-grooved surfaces. Cm, culture medium. (B) Top, AFM topography scan of the CD-R PDMS replica. Blue arrow, groove depth (0.15 μm). Black arrow, ridge width (0.8 μm). Red arrow, groove width (1 μm). Bottom, 10x10 μm scaled surface render. (C) Scanning electron microscope (SEM) image of the CD-R PDMS replica. Scale bar, 5 μm. (D) Left, phase image of HeLa cells confined on the PDMS nano-grooved surface at 3 μm. Scale bar, 10 μm. Right, higher magnification view of the same experiment. Scale bar, 10 μm. (E) Steps in the fabrication of the glass bottom petri dish with PDMS nano-grooved surface.
Fig. 3 Dynamic cell confinement combined with collagen inclusion. (A) Side view of the confiner coverslip setup combined with collagen inclusion. Cs, unpolymerized collagen solution. (B) Representative confocal spinning disk field of view of 3 μm-confined Actin-GFP-expressing HeLa cells embedded in a collagen gel. Cyan, Actin-GFP channel. Grey, Collagen channel. Scale bar, 10 μm. (C) Representative bottom, middle, and top confocal spinning disk images of a 3 μm-confined Actin-GFP HeLa cell embedded in a collagen gel. Cyan, Actin-GFP channel. Grey, Collagen channel. Scale bar, 10 μm.
Fig. 4 Cell fixation under agarose confinement. (A) Left, normal surfaces of the 3D-printed agarose holder. Right, orthogonal view of the 3D-printed agarose holder. (B) Left, the fabrication process of the agarose pad. Ag, agarose solution. Gc, glass coverslip. Pf, parafil...
Fig. 2 Cell confinement combined with nano-grooved surfaces. (A) Side view of the confiner coverslip setup combined with nano-grooved surfaces. Cm, culture medium. (B) Top, AFM topography scan of the CD-R PDMS replica. Blue arrow, groove depth (0.15 μm). Black arrow, ridge width (0.8 μm). Red arrow, groove width (1 μm). Bottom, 10x10 μm scaled surface render. (C) Scanning electron microscope (SEM) image of the CD-R PDMS replica. Scale bar, 5 μm. (D) Left, phase image of HeLa cells confined on the PDMS nano-grooved surface at 3 μm. Scale bar, 10 μm. Right, higher magnification view of the same experiment. Scale bar, 10 μm. (E) Steps in the fabrication of the glass bottom petri dish with PDMS nano-grooved surface.
Fig. 3 Dynamic cell confinement combined with collagen inclusion. (A) Side view of the confiner coverslip setup combined with collagen inclusion. Cs, unpolymerized collagen solution. (B) Representative confocal spinning disk field of view of 3 μm-confined Actin-GFP-expressing HeLa cells embedded in a collagen gel. Cyan, Actin-GFP channel. Grey, Collagen channel. Scale bar, 10 μm. (C) Representative bottom, middle, and top confocal spinning disk images of a 3 μm-confined Actin-GFP HeLa cell embedded in a collagen gel. Cyan, Actin-GFP channel. Grey, Collagen channel. Scale bar, 10 μm.
Fig. 4 Cell fixation under agarose confinement. (A) Left, normal surfaces of the 3D-printed agarose holder. Right, orthogonal view of the 3D-printed agarose holder. (B) Left, the fabrication process of the agarose pad. Ag, agarose solution. Gc, glass coverslip. Pf, parafil...