Methodological approach for the detection of both microdamage and fluorochrome labels in ewe bone and human trabecular bone.

The purpose of this study was to adapt various staining methods for the detection of microdamage in human bone, while preserving tetracycline labels. We describe two staining methods using calcein green and xylenol orange, first developed in ewe bone samples and validated in human trabecular bone samples. In ewe bones, we found that calcein green at 0.5 mM concentration diluted in 100% ethanol as well as xylenol orange at 5 mM were the most adequate fluorochromes both to detect microdamage and preserve the double tetracycline labeling. These results were verified in human trabecular bone (iliac crest for the tetracycline label, and vertebral bone for the double labeling). Results obtained in human bone samples were identical to those in ewes, so this combination of fluorochromes is now used in our laboratory.