Thin poly(vinyl alcohol) cryogels: reactive groups, macropores and translucency in microtiter plate assays

Thin macroporous poly(vinyl alcohol) (PVA) hydrogels were produced by cross-linking of PVA in a semi-frozen state with glutaraldehyde (GA) on glass slides or in the wells of microtiter plates. The 100-130 μm-thick gels were mechanically transferable, squamous translucent films with a high porosity of 7.2 ± 0.3 mL/g dry PVA i.e. similar to larger cylindrical PVA monoliths of the same composition. Additional treatment of the gels with 1% GA increased the aldehyde group content from 0.7 to 2.4 μmol/mL as estimated using dinitrophenylhydrazine (DNPH) reagent. Translucency of the gels allowed registration of UV-visible spectra of the DNPH-stained films. The catalytic activity of trypsin covalently immobilized on thin gels in the microtiter plates was estimated with chromogenic substrate directly in the wells, and indicated that the amount of protein immobilized was at least 0.34 mg/mL gel. Human immunoglobulin G (IgG) immobilized on thin gels at 0.1-10 mg/mL starting concentrations could be detected in a concentration-dependent manner due to recognition by anti-human rabbit IgG conjugated with peroxidase and photometric registration of the enzymatic activity. The results indicate good permeability of the hydrogel pores for macromolecular biospecific reagents and suggest applications of thin reactive PVA hydrogels in photometric analytical techniques.