Sp5 inverted microscope

Hydrogels were imaged by multi-photon second harmonic generation (MP-SHG) in PBS using a Leica SP5 inverted microscope equipped with a MaiTai HP DeepSee multi-photon laser (Spectraphysics) on a 25× NA objective. Second harmonic signal was generated at 900 nm and detected on a photomultiplier tube (PMT) (435–465 nm).

Cross-sectional morphological examination of all foams in a dry state was performed. Foams were imaged by scanning electron microscopy (SEM) using a JEOL 5610 (Herts, UK). Samples were coated with 100 Å Au using an Emitech K550 sputter coater prior to imaging at an accelerating voltage of 15 kV and a working distance of 150 mm.
Foams were incubated in PBS, 0.5 mg mL⁻¹ HA (Creative PEGWorks, UK), or 0.5 mg mL⁻¹ CS (Creative PEGWorks, UK) for 24 h at 37 °C in a 5% CO₂ atmosphere, washed three times in PBS and imaged by multiphoton second harmonic generation (MP–SHG) in wet state using a Leica SP5 inverted microscope equipped with a MaiTai HP DeepSee multiphoton laser (Spectraphysics) on a 25× NA objective. Second harmonic signal was generated at 900 nm and detected on a photomultiplier tube (PMT) (435–465 nm). Collagen samples were further characterized using Fourier transform infrared (FTIR) spectroscopy. FTIR analysis with a PerkinElmer Spectrum One spectrometer was used to determine FTIR spectra showing characteristic peaks for all samples. FTIR spectra were taken with a scanning wavenumber range from 4000 to 650 cm⁻¹.