# Identification and elemental mapping of enamel minerals with electron energy-loss spectroscopy

## TL;DR

This paper introduces a new method using electron energy-loss spectroscopy to distinguish between two similar enamel minerals and apply it to mouse teeth.

## Contribution

The study introduces a novel protocol for using EELS to differentiate HA and OCP in biological specimens.

## Key findings

- The oxygen K-edge in EELS spectra helps distinguish HA from OCP.
- The Ca/P ratio calculated with Kexp provides a second indicator for mineral differentiation.
- EELS analysis was successfully applied to study enamel mineral development in mice.

## Abstract

The identification and differentiation of the mineral phases, hydroxyapatite (HA, Ca10(PO4)6(OH)2) and octacalcium phosphate (OCP, Ca8H2(PO4)6), remains challenging because of their similar composition and chemical structure. In this research, electron energy-loss spectroscopy (EELS) analyses revealed indicators to distinguish HA and OCP and these were applied to examine mineral development in enamel from mouse incisors. Reference EELS data for HA and OCP was established with commercial HA and synthesized OCP. An evaluation of electron damage and a mitigation strategy of multipass imaging was conducted, and the electron dose limitation of OCP was identified. New insights into the mechanism of electron beam damage on the apatite crystal were obtained. With the characterization of the energy-loss spectra and the EELS simulation, the oxygen K-edge was found to be one of the indicators for the differentiation of HA from OCP. The second indicator, the Ca/P ratio, was calculated with a calibrated experimental factor of Kexp. Elemental mapping was done to establish the different Ca/P ratio of HA and OCP, and the boundary between these mineral forms. EELS analysis was performed on developing enamel in wild-type (WT) and Mmp20 knockout (KO) mice. This research establishes a protocol for EELS analysis on biological specimens and demonstrates the power and potential of EELS in biomaterial characterization.

Investigation and characterization of mineral phases in dental enamel using electron energy loss spectroscopy.