Immunofluorescent Staining
Haley Geertsma
Abstract
This protocol is used to stain cryosectioned mouse brain tissue.
Steps
To cryo-sectioned brain tissue, wash with 1X phosphate buffered saline (PBS) for 3x 5-minute washes.
Incubate in blocking buffer for 1 hour at room temperature.
Blocking buffer: 10% serum + 0.5% Triton X-100 in 1X PBS
Wash tissue with 1X PBS.
Incubate in primary antibody diluted in blocking buffer overnight at 4oC.
Wash tissue with 1X PBS for 5x 5-minute washes.
Incubate in secondary antibody diluted in blocking buffer for 1 hour at room temperature.
Wash tissue with 1X PBS for 5x 5-minute washes.
If tissue wasn't previously mounted on a slide, mount on a superfrost plus slide and let dry at room temperature for at least 10 minutes.
Coverslip with fluorescent mounting medium and a #1.5 coverslip. Outline the coverslip with clear nailpolish.
