Rapid bone staining with hair removal (RAP-B/HR): a non-destructive and rapid whole-mount bone staining protocol optimized for adult hairy mice

Abstract

We developed a non-destructive and rapid whole-mount bone staining method for small fish, Xenopus laevis, and rodent fetuses (RAP-B). RAP-B does not require skin or soft tissue removal. However, RAP-B requires hair removal from hairy animals, such as adult mice and rats. In the present study, we investigated hair removal chemical treatments that did not result in soft tissue destruction. The hair removal effectiveness was investigated using a calcium mercaptoacetate or sodium mercaptoacetate solution on skin fragments obtained from the back of adult mice. A mixture of 2% sodium mercaptoacetate in 3% potassium hydroxide was found to be the most effective in complete hair removal from the skin. Using this hair removal treatment as a pretreatment for RAP-B, the preparation of fast-acting artifact-free whole-mount bone staining was possible without skin and soft tissue removal (RAP-B/HR). We performed a seamless observation from a low magnification wide-view to a high magnification without artifactacting artifacts using fluorescence zoom microscopy. Therefore, the combination of RAP-B/HR and fluorescent zoom microscopy is a novel platform for three-dimensional, wide-field, high-resolution pathological anatomical analysis.