Transfection Techniques and Combined Immunocytochemistry in Cell Cultures and Organotypic Slices

Transfection techniques in vitro and ex vivo (organotypic cultures ) offer an array of possibilities to investigate the consequence(s) of the introduction of foreign nucleic acids (DNA but also RNA ) and or other biologically active molecules into neurons , and to combine observations with immunocytochemistry . In particular, a wide number of fluorescent reporter proteins (FRPs ) can be employed for multicolor fluorescence imaging . Here we present a series of protocols for in vitro and ex vivo transfection of DNA or RNA sequences into cerebellar neuron cultures and organotypic slices based on the use of plasmid vectors and multicolor laser scanning confocal microscopy . These protocols allow analysis of live transfected cells , and, after fixation, correlative neurochemical studies.