Materials
PdBT synthesis
Preparation of azide-presenting biomolecules
Click functionalization of PdBT
1H NMR spectroscopy
GPC determination of molecular mass
Biofunctionalized hydrogel fabrication
Differential scanning calorimetry of cross-linking kinetics
Colorimetric assessment of biomolecule incorporation in hydrogels
Assessment of biofunctionalized hydrogel degradation
Leachable cytotoxicity assay
MSC harvest and culture
In vitro MSC encapsulation
Statistical analysis
The ability of PdBT, CS/PdBT, BMPm/PdBT, and NC/PdBT to cross-link P(NIPAAm-co-GMA) was assessed by a study in which fabricated hydrogels were swollen in excess PBS and then assessed for degree of mass swelling, i.e., the ability to resist the natural tendency of P(NIPAAm-co-GMA) to shrink and expel water mass after formation (3). In this study, hydrogels were weighed immediately after fabrication (Wf) and then weighed again after equilibrium swelling (Ws) in 1.5 ml of PBS at pH 7.4 for 24 hours at 37°C. Swollen hydrogels were then flash-frozen in liquid N2 and lyophilized to obtain the dry weight (Wd). From these weights, the initial formation swelling ratio was calculated as Wf−WdWd, while the equilibrium swelling ratio was calculated as Ws−WdWd. The minimum PdBT, CS/PdBT, BMPm/PdBT, and NC/PdBT concentrations that resulted in a greater equilibrium swelling ratio than the initial formation swelling ratio, indicating sufficiently cross-linked systems, were then used for all further hydrogel characterization.