Traditionally, cultures of primary cortical neurons are prepared from embryonic animals because at prenatal stages neurons have not yet developed extensive axonal and dendritic arbors and are not highly innervated, thus rendering the cells less susceptible to damage during dissociation of the neuronal tissue. The appropriate developmental age for preparing primary cultures of any cell type is determined by the time at which the cells of interest are generated and abundant. Most cerebral cortical neurons are generated between embryonic days (E) 11 and 17 in the mouse (embryos being considered 0.5 days old when a vaginal plug is detected in the morning). Here we describe a method to obtain short-term cultures of mouse primary cortical neurons at E15.5 and a practical application using fluorescent immunocytochemistry.