Acid-etching technique of non-decalcified bone samples for visualizing osteocyte-lacuno-canalicular network using scanning electron microscope

The aim of this study was to define the acid-etching technique for bone samples embedded in polymethyl metacrylate (PMMA) in order to visualize the osteocyte lacuno-canalicular network (LCN) for scanning electron microscopy (SEM). Human jaw bone tissue samples (N = 18) were collected from the study population consisting of patients having received dental implant surgery. After collection, the bone samples were fixed in 70% ethanol and non-decalcified samples embedded routinely into polymethyl metacrylate (PMMA). The PMMA embedded specimens were acid-etched in either 9 or 37% phosphoric acid (PA) and prepared for SEM for further analysis. PMMA embedded bone specimens acid-etched by 9% PA concentration accomplishes the most informative and favorable visualization of the LCN to be observed by SEM. Etching of PMMA embedded specimens is recommendable to start with 30 s or 40 s etching duration in order to find the proper etching duration for the samples examined. Visualizing osteocytes and LCN provides a tool to study bone structure that reflects changes in bone metabolism and diseases related to bone tissue. By proper etching protocol of non-decalcified and using scanning electron microscope it is possible to visualize the morphology of osteocytes and the network supporting vitality of bone tissue.