Spatial quantification of hydrogels swelling using wide-field fluorescence microscopy

Wide-field fluorescence microscopy (WFM) is used to spatially quantify the protein content of large hydrogels during swelling. Whey protein gels made at different protein concentrations, labelled with rhodamine B isothiocyanate (RITC), were used as a model system. Labelling and swelling measurement conditions were optimized. Dynamic swelling experiments at different pH agree very well with the expected fluorescence decrease for isotropic gels using overall volumetric data, despite the existence of internal gradients. Deviations are observed at large swelling degrees, provably due to protein-dye leakage, and at high protein concentrations. This simple and ubiquitous technique is used to spatially quantify the swelling of protein hydrogels in 2D at different swelling times, highlighting the existence of a variety of swelling profiles inside the gels with time.