2.2.2. Characterization of Hydrogel-Forming MN Arrays

Following fabrication, and prior to use in both in vitro and in vivo studies, fabricated MN arrays were viewed under a digital light microscope (Leica Microsystems, Milton Keynes, Buckinghamshire, U.K.) with the height of individual MNs measured and recorded as H_b.

To test the insertion efficiency of the formulated MN arrays, a TA.XT.Plus Texture Analyser (Stable MicroSystems Ltd., Godalming, Surrey, U.K.) and an artificial skin model composed of eight layers of Parafilm M were used according to the widely accepted protocol previously reported by Larrañeta et al.¹⁸ Insertion efficiency was calculated using eq 1

Additionally, MN height reduction after insertion into the artificial Parafilm M skin model was carried out by visualizing MNs post-insertion using a Leica EZ4 D digital microscope, measuring individual MN heights (H_a), and comparing these with MN heights before insertion (H_b) using eq 2

The insertion depth of MNs into full-thickness neonatal porcine skin was assessed using the same insertion protocol as previously. An OCT microscope was then used to obtain cross-sectional images of the inserted MNs, and ImageJ software (National Institutes of Health, Bethesda, MD) was used to process the obtained images.

Determination of the swelling profile of each hydrogel formulation was conducted using an excess volume of phosphate-buffered saline (PBS) (pH 7.4), as described previously by Donnelly et al., (2014).¹⁶ The percentage swelling of each film was calculated using eq 3, where M₀ represents the starting mass of the hydrogel film and M_t represents the mass of the swollen hydrogel film at a given timepoint