Recipes/Live Alizarin bone stain

From Kimmel Lab Wiki

30 January 2009

LIVE BONE STAIN with Alizarin

Alizarin Red S (sodium alizarin sulfonate):

• Stock solution is 0.5g/100ml sterile water.
• Final: 50 μg/ml.

• Use 10μl stock solution per 1 ml embryo medium.
• Buffer w/ 10 mM HEPES.

Recipes (add in order to prevent precipitation):

For 1 Liter:

• 10 ml 1M HEPES
• 980ml Embryo medium

• Mix.

• Add 10 ml 0.5% Alizarin Stock in sterile water to get 1000 ml total

For one 100mm diameter petri dish:

Mix in 50 µl culture tube.

• 500 µl 1 M HEPES
• 49 ml Embryo medium

• Add 500 µl 0.5% Alizarin Stock in sterile water to get 50 ml total

Stain overnight in the dark.

Rinse thoroughly with embryo medium.

Can observe right away, but excess stain in tissue continues to go down with time.

Bone fluorescence will decrease over time, so plan on imaging the same day if possible.

Keep fish in the dark as much as is reasonably convenient.