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Nissl staining

AK Amy Keerie
HB Heledd Brown-Wright
IK Isaac Kirkland
AG Andrew Grierson
JA James J. P. Alix
CH Christian Holscher
RM Richard J. Mead
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Rehydrated sections were incubated in a 0.1% cresyl violet solution for 15 min. Slides were then rinsed in H2O and incubated in acetic acid for 4–8 s. Slides were then dehydrated in 100% ethanol for 1 min and cleared in 100% xylene prior mounting with DPX mountant. Images were collected using a nanozoomer digital slide scanner (Hamamatsu) and analysed in NDP.view 2 software (Hamamatsu). Motor neurons were then counted using a criteria of > 20 μm in cell body size in any axis, location I the ventral horn and presence of a nucleus and nucleolus. Counting of motor neurons was conducted blinded.

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