Sequential labelling of microdamage in bone using chelating agents

Basic fuchsin labels microcracks, but a series of stains is required to differentiate between preexisting and test‐induced microcracks and to label their growth in vitro. Basic fuchsin and five chelating agents—alizarin complexone, calcein, calcein blue, oxytetracycline, and xylenol orange—were randomly assigned to label microcracks in sequential rib sections from 10 donors. The density, length, and location of the microcracks did not differ significantly between he six stains, suggesting that each was equally effective in detecting microcracks. Paired specimens of trabecular bone were machined from bovine tibiae, stained with oxytetracycline, and fatigued in compression. One specimen from each pair was then stained with xylenol orange. Preexisting microdamage was stained with oxytetracycline, propagating microcracks with both stains and new, test‐initiated damage with xylenol orange. Chelating agents are site‐specific markers of the initiation and growth of microcracks.