Alcian Blue Staining: Difference between revisions
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(Created page with "# Fix sample O/N in 4% PFA freshly made # Wash in PBS, 3X5mins, 3X30mins 3. Wash 20mins in PBST 4. Equilibrate in EtOH, 25, 50%, 10 mins each 5. Place in Alcian blue staini...") |
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# Fix sample O/N in 4% PFA freshly made | # Fix sample O/N in 4% PFA freshly made | ||
# Wash in PBS, 3X5mins, 3X30mins | # Wash in PBS, 3X5mins, 3X30mins | ||
# Wash 20mins in PBST | |||
# Equilibrate in EtOH, 25, 50%, 10 mins each | |||
# Place in Alcian blue staining solution: 9mg alcian blue dissolved in 60mls EtOH plus 40mls glacial acetic acid , make fresh every time. | |||
# Staining is carried out at 37C, can take anytime between 1 hr to 2days. Check every so often as the solution will precipitate. | |||
# When staining is complete wash with EtOH/acetic acid mix for 1hr at RT | |||
# Rehydate gradually, 90, 70, 50 25% EtOH/PBS. | |||
# Then go through glycerol series 25, 50, 75% glycerol/PBS | |||
Revision as of 08:40, 13 February 2015
- Fix sample O/N in 4% PFA freshly made
- Wash in PBS, 3X5mins, 3X30mins
- Wash 20mins in PBST
- Equilibrate in EtOH, 25, 50%, 10 mins each
- Place in Alcian blue staining solution: 9mg alcian blue dissolved in 60mls EtOH plus 40mls glacial acetic acid , make fresh every time.
- Staining is carried out at 37C, can take anytime between 1 hr to 2days. Check every so often as the solution will precipitate.
- When staining is complete wash with EtOH/acetic acid mix for 1hr at RT
- Rehydate gradually, 90, 70, 50 25% EtOH/PBS.
- Then go through glycerol series 25, 50, 75% glycerol/PBS