Abstract:
OBJECTIVE : Retinoic acid (RA) is known to transition proliferating SH-SY5Y neuroblastoma cells towards functional
neurons. However, the activity of RA is restricted due to its photolability where any findings from prolonged time
course observations using microscopy may alter outcomes. The aim of the study was to establish a real-time, longterm
(9-day) protocol for the screening of differentiation events using Electrical cell-substrate impedance sensing
(ECIS).
RESULTS AND DISCUSSION : A differentiation baseline for SH-SY5Y cells was established. Cells were seeded and exposed
to repeated spikes of RA using the xCELLigence real-time cell analyser single plate (RTCA-SP) for real-time monitoring
and identification of differentiation activity over a 9 day period in order to be more representative of differentiation
over a prolonged timeline. Specific features associated with differentiation (growth inhibition, neurite outgrowths)
were confirmed by end-point analysis.
RA-induced growth inhibition and assumed phenotypic changes (i.e. neurite outgrowth) were identified by the
xCELLigence analysis and further confirmed by end-point metabolic and phenotypic assays. Change in cellular
morphology and neurite outgrowth length was identified by end-point fluorescence detection followed by
computational analysis. Based on this it was possible to identify SH-SY5Y phenotypic differentiation with distinct
phases observed over 9 days using Electric cell-substrate impedance sensing (ECIS) cell index traces providing a path
to application in larger scale neurotrophic factor screening using this scalable technology.
