I noticed in the brains I've cleared there has been a white precipitate forming when placed in Focus Clear, and when viewed under the microscope there are very noticeable holes in the tissue. We think it may be the result of the way we made the clearing solution. When making the solution, we just added the boric acid and sds, followed by the dH2O. A white precipitate formed, and it took more than 200 pellets of NaOH (for 10L of clearing solution) to bring the pH to 8.5.
Has anyone had this issue, or is there a specific protocol that works better for making the clearing solution (like dissolving either boric acid or sds first?). Thanks!
I will recommend first prepare the boric acid and adjust pH, when the pH is
set to 8.5 add the SDS. At the end check the pH again with some pH paper
Still, I agree with Fabio's method to prepare the clearing solution. I usually recommend preparing a 2L solution of boric acid in water and adjusting the pH to 8.5 (you will probably need to start adding NaOH to get the boric acid to dissolve completely). You can then add the 2L solution to a 10L container containing the SDS and add the necessary volume of water. Check the pH, but it should remain stable at 8.5.
Instead of storing our samples in FocusClear, we keep them in PBS until the day before we are ready to image, and then place them in FocusClear.
1) A distributor of Focus Clear in Europe. Apparently it looks like cell explorer labs is from taiwan. Any suggestions for focus clear distributors in Denmark?
2) Does anybody has a experience of imaging the ETC cleared tissue under BABB (Murray's Clear) ?