The tooth, the whole tooth and nothing but the tooth….

 Tara Nylese, Global Applications Manager

Tooth enamel is the hardest material in the human body. While it is easy to think biological materials are not a very interesting target for x-ray microanalysis, there are often unknowns lurking in the small and difficult to reach places. This is the case with a cavity just forming in a tooth.

When my 10 year old daughter lost her tooth last year, I was intrigued by the ‘small’ brown spot that I noticed at the base of the crown of the tooth. Since small is a relative term in the world of microscopy and microanalysis and there’s always a hunt for new and interesting samples, I traded with the tooth fairy and gained a new sample.

Despite the irregular shape of this premolar, I was easily able to mount it on a holder with carbon tape and I then brought it into the SEM under low vac mode to help conduct away the electron beam charge from the insulating material. Working at low vac is important for biological materials since introducing carbon coating for charge reduction creates confusion in native carbon distribution, and heavier metal coatings absorb the more delicate low energy x-rays which must escape the tissue. I was quickly able to see morphological differences in the suspected cavity area, with fine cracks and a sunken in appearance that wasn’t visible to the eye. I then used one of the primary tools in a microanalyst’s toolkit, BSE imaging, to see atomic number contrast in the image, indicating gross changes in the chemical makeup around the sunken area.

Image showing morphological differences and atomic number contrast

Image showing morphological differences and atomic number contrast

Within seconds of collecting the image, I gathered a spectrum from an area of less than one millimeter across.  At moderate beam current, which the sample handled easily, the overall chemistry was immediately apparent with the clearly recognizable O, P and Ca peaks, even before the EXpert ID kicked in five seconds later.

Spectrum showing O, P and Ca peaks

Spectrum showing O, P and Ca peaks

As anticipated, this is consistent with this being hydroxyapatite (HA), which is the main mineral component of tooth enamel and bones.

The main reason for the formation of dental cavities is the erosion of the enamel by acid from eating sugars; erosion then permits bacterial invasion within this area.  After the first spectrum collection, it was just a matter of tracking the chemical distribution around the cracks and with a few survey spectra, the carbon content was found to be highest in the darker areas of the BSE image.  The presence of carbon indicates organic matter in the form of a bacterial biofilm within the eroded area.

Since visual distribution is the surest way to understand the material, the next step was to collect an x-ray map.  During the collection I selected the major constituents and dynamically overlaid them to understand how the chemistry matched up with the morphology, Ca and C shown here.

Ca and C overlay map

Ca and C overlay map

The cracks and holes are confirmed with the unique CPS map, which shows variations of x-ray intensities and lends a better understanding of not only elemental intensities, but also the impact of topography on the appearance of those elements.

Unique counts per second (CPS) amp which confirms cracks and holes

Unique counts per second (CPS) map confirms cracks and holes

Along with this function there is a normalization routine, which is applied to make up for the reduction of counts in the cracks and holes particularly for elements that are truly present throughout, such as can be seen with the Oxygen map before and after maps here.

Oxygen map before normalization routine Oxygen map after normalization routine
Oxygen map before normalization routine  Oxygen map after normalization routine 

A final note here is that, interestingly, it is not directly the amount of sugar at one time that leads to the enamel loss but extended exposure throughout the course of the day.  Better to eat one’s Easter basket in one sitting than making it last throughout the day to avoid cavities!

 

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