RESEARCH ARTICLE


A Method for Rapid Demineralization of Teeth and Bones



Andrew Cho1, Shigeki Suzuki1, Junko Hatakeyama1, Naoto Haruyama1, Ashok B Kulkarni1, 2, *
1 Gene Targeting Facility, Laboratory of Cell and Developmental Biology, National Institute of Dental and Craniofacial Research, National Institutes of Health, 30 Convent Drive, Bethesda, MD 20892, USA
2 Functional Genomics Section, Laboratory of Cell and Developmental Biology, National Institute of Dental and Craniofacial Research, National Institutes of Health, 30 Convent Drive, Bethesda, MD 20892, USA


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Creative Commons License
© Choet al.; Licensee Bentham Open.

open-access license: This is an open access article licensed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted, non-commercial use, distribution and reproduction in any medium, provided the work is properly cited.

* AAddress correspondence to this author at the Chief, FGS, CDBRB, NIDCR, NIH, Tel: 301-435-2887; Fax: 301-435-288; E-mail: ak40m@nih.gov


Abstract

Tooth and bone specimen require extensive demineralization for careful analysis of cell morphology, as well as gene and protein expression levels. The LacZ gene, which encodes the ß-galactosidase enzyme, is often used as a reporter gene to study gene-structure function, tissue-specific expression by a promoter, cell lineage and fate. This reporter gene is particularly useful for analyzing the spatial and temporal gene expression pattern, by expressing the LacZ gene under the control of a promoter of interest. To analyze LacZ activity, and the expression of other genes and their protein products in teeth and bones, it is necessary to carry out a complete demineralization of the specimen before cutting sections. However, strong acids, such as formic acid used for tooth demineralization, destroy the activities of enzymes including those of ß-galactosidase. Therefore, most protocols currently use mild acids such as 0.1 M ethylene diamine tetra-acetic acid (EDTA) for demineralization of tooth and bone specimen, which require a longer period of treatment for complete demineralization. A method by which hard tissue specimens such as teeth and bones can be rapidly, but gently, decalcified is necessary to save time and effort. Here, we report a suitable method for rapid demineralization of mouse teeth in 0.1M EDTA at 42˚C without any loss of ß-galactosidase activity.

Key Words: Demineralization, tooth, LacZ, gene expression.