Localized functional chemical stimulation of TE 671 cells cultured on nanoporous membrane by calcein and acetylcholine

Abstract

Acetylcholine sensitive TE 671 cells were cultured on nanoporous membranes and chemically stimulated by localized application of i), calcein-AM and ii), acetylcholine, respectively, onto the bottom face of the membrane employing an ink jet print head. Stimulus correlated response of cells was recorded by fluorescence microscopy with temporal and spatial resolution. Calcein fluorescence develops as a result of intracellular enzymatic conversion of calcein-AM, whereas Ca2+ imaging using fluo-4 dye was employed to visualize cellular response to acetylcholine stimulation. Using 25 pl droplets and substance concentration ranging from 10 μM to 1 mM on Nucleopore membranes with pore diameters between 50 nm and 1 μm, a resolution on the order of 50 μm was achieved. © 2007 by the Biophysical Society.