A Novel and Robust Protocol for Differentiation of SH-SY5Y Neuroblastoma Cells into Neuron Like Cells

Human neuroblastoma cell line SH-SY5Y is a frequently used experimental cellular model in a variety of neuropsychiatric and neurodegenerative disorders. It is crucial to use a culture protocol that supports the fully differentiation of SH-SY5Y into neuron-like phenotype for the consistency of the results with neurons in vivo. However, a standardized neuronal differentiation protocol for SH-SY5Y cells still does not exist. Numerous differentiation methods have been proposed in the literature, yet SH-SY5Y cells with stronger neuronal characteristics and a more favorable environment for these differentiated cells are required in order to best representation of neurons. Therefore, in the study, we aimed to establish a more successful differentiation protocol for SH-SY5Y cells based on the primary neuron culture technique, which neuronal maturation is very well defined.

In the study, we rearranged previous SH-SY5Y differentiation protocols, combined them with our primary neuron culture protocol and created a robust and reproducible protocol for differentiation of SH-SY5Y.

Our proposed "retinoic acid+brain-derived neurotrophic factor (RA+BDNF)-induced 7 days differentiation (conalbumin- on day 4) protocol provided well developed neurites, adequate expression and localization of neuronal and synaptic markers resembling mature neurons.

The differentiation protocol we present can enable researchers to obtain satisfactory and properly differentiated SH-SY5Y cells in each independent experiment, achieving the closest possible in vivo results.