Undecalcified bone histology is a valuable diagnostic method for studying bone microarchitecture and provides information on bone formation, resorption, and turnover. It has various clinical and research applications. Toluidine blue has been widely adopted as a staining technique for hard-tissue specimens. It provides a clear identification of bone structural and cellular features and the distinctions between them. Furthermore, the method allows for an excellent definition of the cement lines that mark the fields of bone remodeling. Some of the suggested and currently used processing and staining protocols are too complex and time-consuming, which necessitates their modification and/or optimization. This research aims to develop a simplified protocol for staining plastic-embedded undecalcified bone specimens with toluidine blue. The samples were obtained from the tibial bones of rabbits, and experiments with and without pre-etching were conducted. Our results demonstrated that the optimal visualization of the bone microstructure and its cellular components was achieved in the samples without acid pre-etching and dehydration after staining.

Bone histology and histomorphometry are reliable diagnostic tools for the assessment of the bone–implant interface, material safety and biocompatibility, and tissue response. They allow for the qualitative and quantitative analysis of undecalcified bone specimens. This scoping review aims to identify the most common staining techniques, study models for in vivo experiments, and histomorphometric parameters used for quantitative bone evaluation of osseointegrated dental implants in the last decade. The Web of Science, PubMed, and Scopus databases were searched on 1 July 2024 for relevant articles in English, published in the last ten years, and the data were exported to an MS Excel spreadsheet. A total of 115 studies met the eligibility criteria and were included in the present review. The results indicate that the most common study models are dogs, rabbits, and pigs. Some of the most frequently used methods for the assessment of the bone–implant interface are the Toluidine blue, Stevenel’s blue with Van Gieson, and Levai–Laczko stainings. The results from this study demonstrate that the most commonly used histomorphometric parameters in implant dentistry are the bone-to-implant contact (BIC), bone area fraction occupancy (BAFO), bone area (BA), and bone density (BD). This review presents the recent trends in histomorphometric analysis of dental implants and identifies some research gaps that necessitate further research.

With the implementation of bone substitute materials, regeneration strategies have inevitably evolved over the years. Histomorphometry is the optimal means of quantitative evaluation of bone structure and morphology. This systematic review focuses on determining study models, staining methods and histomorphometric parameters used for bone regeneration research on non-decalcified plastic-embedded specimens over the last 10 years. After being subjected to the inclusion and exclusion criteria, 118 studies were included in this review. The results establish the most commonly selected animal model is rat, followed by rabbit, sheep and dog. Strong preference for staining samples with toluidine blue was noted. With regard to histomorphometric parameters, terms related to bone were most frequently assessed, amounting to almost half of recorded parameters. New bone formation was the main descriptor of this category. Residual bone graft and non-bone tissue parameters were also often evaluated. With regard to dynamic histomorphometry, mineral apposition rate (MAR) was the parameter of choice for most researchers, with calcein green being the preferred dye for fluorochrome labelling. An overview of the contemporary literature, as well as weaknesses in the current research protocols have been discussed.