Calcein Blue and Tetracycline Bone Fluorescence

From:Ryan Blair Rountree

I am trying to characterize the bone deposition rates in mouse feet.  I
have read several papers and plan to inject calcein blue first, wait 7
days, inject tetracycline, wait 2 days, then plastic embed and section.
However, I still have a few questions and any tips would be greatly

What dosage is best for calcein blue?  I have seen it used at 32mg/kg, but
this was for sacrificing the animals 6 hours later.  I plan to sacrifice
the animal 9 days later - should I use a lower dose?

What concentration and in what (water?) do you make up the calcein blue
and tet STOCK solutions? How do you store them?

How careful do you have to be during the sectioning to prevent light
exposure?  Do you have to put the slides immediately into a dark box as
you cut, or can you wait until the session is done?

Will a filter usually used for detecting the DNA stain DAPI that lets
360-380nm light through excite tetracycline enough to visualize it? It's
excitation peak is 390nm (range 380-420), and this borderline.

Thanks for your help!


Ryan Rountree
Department of Developmental Biology
Stanford, CA 94305
Fax: (650) 725-7739

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