title: Characterization of 3-D collagen hydrogels for functional cell-based biosensing.
abstract: To address the growing demand for functional cell-based assay technologies with accelerated drug discovery applications, we have proposed the use of human neuroblastoma cells (IMR-32) immobilized in three-dimensional (3-D) collagen hydrogel matrices. The gel protects weakly adherent cells from fluid mechanical forces while providing a more physiologically relevant 3-D environment. Hydrogels made up of collagen, between 0.5 and 1.0mg/ml, exhibited mechanical stability adequate to withstand fluid mechanical forces (<0.11 mN) typical of automated commercial fluid transfer equipment. Collagen-entrapped cells visualized with the aid of confocal microscopy and a potentiometric-sensitive dye, TMRM, exhibited round morphology in comparison to flat morphology typical of cells in two-dimensional (2-D) monolayer cultures. Morphological differentiation characterized by neurite extension and cell aggregation was observed for both 2-D and 3-D cultures. Differentiated IMR-32 cells failed to develop a resting membrane potential typical of excitable cells. Free intracellular calcium was monitored with Calcium Green-1. Depolarization-induced Ca 2+influx was only observed with differentiated 3-D cells unlike 2-D cells, where calcium flux was observed in both differentiated and undifferentiated cells. Taken together, the results revealed that collagen hydrogels (0.5 mg/ml collagen) were suitable structural supports for weakly adherent cells. However, for voltage-dependent calcium channel function applications, further investigations are needed to explain the difference between 2-D monolayer and 3-D collagen-entrapped cells.
authors.affiliation: Cellular Bioengineering Laboratory, Faculty of Engineering, University of Georgia, Athens, GA 30602, USA.
authors.forename: Chen
authors.initials: C
authors.lastname: Mao
authors.forename: William S
authors.initials: WS
authors.lastname: Kisaalita
mesh_terms: D001672:Biocompatible Materials; D015374:Biosensing Techniques; D003094:Collagen; D005109:Extracellular Matrix; D006801:Humans; D020100:Hydrogels; D008422:Materials Testing; D008564:Membrane Potentials; D018613:Microscopy, Confocal; D008858:Microscopy, Phase-Contrast; D018408:Patch-Clamp Techniques
chemical_list: D001672:Biocompatible Materials; D020100:Hydrogels; D003094:Collagen