Omap anti rb hrp

Egr1cKO mice and controls were rapidly and deeply anesthetized with isoflurane (Vet One, cat# 502017) and perfused transcardially with 10% formalin into the left ventricle. The right ventricle was opened to allow for exsanguination. After cardiac arrest was confirmed, the mice went through cervical dislocation. The brain was exposed with maximum bone removed to allow for further tissue fixation in 10% formalin overnight. Once fixed, tissues were sent to the Virginia-Maryland College of Veterinary Medicine pathology laboratory for embedment. Coronal slices were taken of the brain near the hippocampus in the wax block and provided back to the pathology laboratory. The coronal brain sections went through an automotive system with a Ventana Discovery Ultra machine (DAB Detection kit Cat#: 760–159) and secondary antibody, Omap anti-Rb HRP (Roche, Cat#: 760–4,311). The brain section was stained with anti-EGR1 rabbit antibody (CST, cat# 41542). Images were acquired using a MoticEasy Scan Pro 6, slide scanner.

The Egr1cKO mice and the controls were rapidly and deeply anesthetized with isoflurane (Vet One, Boise, ID, USA, cat# 502017) and perfused transcardially with 10% formalin into the left ventricle. The right ventricle was opened to allow for exsanguination. After cardiac arrest was confirmed, the mice went through cervical dislocation. The brain was exposed and the maximum amount of bone was removed to allow for further tissue fixation in 10% formalin overnight. The pituitary was left in the sella turcica and dissected away from the sphenoid bone. It was then fixed for 48 h in 10% formalin before being removed. Both tissues, once fixed, were sent to the pathology laboratory of the Virginia–Maryland College of Veterinary Medicine for embedment. Coronal slices of the tissues in the wax block were taken and sent back to the pathology laboratory. Both the whole brain and the pituitary gland went through an automotive system involving a Ventana Discovery Ultra machine (DAB Detection kit Cat#: 760-159) and secondary antibody OMap anti-Rb HRP Cat#: 760-4311, purchased from Roche Diagnostic, Indianapolis, USA. Both the whole brain and the pituitary gland were stained with anti-Egr1 rabbit antibody (41542, purchased from Cell Signaling Technology, Danvers, MA, USA). Images were acquired using a MoticEasy Scan Pro 6 slide scanner.