/
Cresyl Violet staining

Cresyl Violet staining

Sep 14, 2022

 

Perform this procedure under a chemical fume hood. Wear two layers of nitrile gloves. Lab coat and eye protection required.

Hazardous chemicals used in this procedure: acetic acid, xylene, DPX

  1. In 0.9% NaCl (aq), spread out vibratome sections of the perfusion-fixed brain in sequence from rostoral to caudal (refer to the rat brain atlas).

  2. Mount the sections in sequence onto glass microscope slides.

  3. Air dry the mounted sections 2-3 days.

  4. Load staining rack with slides with the sections.

  5. Prepare staining dishes: purified water, Cresyl Violet, 100% ethanol, 100% ethanol with 0.1% acetic acid (glacial), 100% ethanol × 4, xylene × 3

  6. Place the rack containing slides in purified water, 2 min

  7. 100% ethanol × 2, 3 min each

  8. Xylene, 4 min

  9. 100% ethanol × 2, 4 min each

  10. 95% ethanol, 4 min

  11. 75% ethanol, 4 min

  12. Purified water × 2, 1 min each

  13. Drain and briefly dry the sections

  14. cresyl violet solution, 2 min

  15. Rise briefly in 100% ethanol

  16. 100% ethanol with 0.1% acetic acid (glacial), ~1-2 min

    1. 200 ml working vol EtOH, 200 µl Acetic Acid                                                     

  17. 100% ethanol × 4, 2 min each (check staining with a microscope after 2 changes)

  18. Xylene × 3, 3 min each

  19. Coverslip with DPX mountant - keep slides in xylene until coverslip is applied

  20. List of slides:

  21. Let dry in hood ~48 hrs