In vivo skull optical clearing technique

In addition, an innovative optical clearing skull window was employed to noninvasively monitor cortical vessels via treatment with a skull optical clearing agent, which consisted of two solutions (solution-1 and solution-2). Solution-1 was a saturated supernatant solution consisting of 75% (vol/vol) ethanol and urea at 25 °C and was applied to the exposed skull of mice for 10 min to make the skull transparent. Solution-2 is a high-concentration sodium dodecyl benzene sulfonate solution that was prepared by mixing 0.7 M NaOH solution with dodecyl benzene sulfonic acid at a volume-mass ratio of 24:5. Solution-2 was applied after solution-1 to enhance skin clearing and was applied for 5 min. The detailed protocol describing our use of these in vivo skull optical clearing agents is provide in our recent study ²⁸.