Cresyl Violet Staining

Paraffin embedded sections were deparaffinized for 45 min at 60C and 2 10 min in Xylene. The sections were sequentially rehydrated for 2 5 min in 99% EtOH, 3 min in 96% EtOH, 3 min in 70% EtOH, rinsed in tap water and stained for 12 min in 37C Cresyl Violet (Millipore Sigma). The stained sections were rinsed in distilled water and staining outside of perikarya was differentiated for 10 min in 96% EtOH with 0.02% glacial acetic acid (Millipore Sigma). The sections were subsequently dehydrated 2 5 min in 99% EtOH and 2 5 min in Xylene prior to coverslip mounting using DPX (Sigma-Aldrich). All images were acquired on an Axio Scan.Z1 (Zeiss) using automated brightfield imaging.