A New Analytical Method for Calcium Quantification in Danio rerio Scales

The scale of Danio rerio is an ossified tissue which can be analyzed histologically, biochemically and immunohistochemically. In this paper, we describe and characterize the validation of a new colorimetric procedure for the quantification of the calcified component of the fish scale. This method is based on the use of alizarin red S (ARS) that specifically binds calcium of the bone matrix and ethylenediaminetetraacetic acid (EDTA), a chelating agent that competes with ARS for the binding of calcium. The release of ARS from the scale can be quantified by measuring the absorbance of the alizarin red contained in the EDTA extraction solution since it is proportional to scale calcium content, then it can be considered an index of the degree of mineralization of the scale itself. This new method has been proved to be accurate, precise, based on linear regression and allows a great characterization of scale mineral content. Then, it can be very helpful to evaluate the mineral alterations induced by different pathologies in bone metabolism studies.