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Optical measurements to determine the thickness of calcite crystals and the mass of thin carbonate particles such as coccoliths


We describe a procedure for measuring the thickness and mass of calcite particles that works for most calcite particles <4.5-μm thick. The calcite particles are observed in cross-polarized light, which enables the light transmitted through the calcite particles to be correlated with their thickness. Three polarizing planes are used to minimize the darkening of crystals at some orientations (black cross). This allows direct measurement of the thickness without recourse to a transfer function. This procedure has been used recently to determine the degree of calcification of coccoliths, which provides an indicator of ocean acidification. It takes only a few minutes per sample, and it is an improvement over the former protocol, which did not allow measurement of the thickness and mass of particles thicker than 1.5 μm.

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Figure 1: Relationship between the thickness of a calcite crystal and the amount of transmitted light.
Figure 2: Images of a rhabdolith.
Figure 3: Three original images of a placolith from left to right: 0°, 35° and 45°, and its merged image on the right.
Figure 4: Relation between hue angle (H° in 8 bits) and retardation (nm).
Figure 5: Effect of focus on measurements of thickness.
Figure 6: Schematic representation of the potential effect of the inclination of a coccolith.
Figure 7: Width (diameter) of rhabdolith stems measured with a microscopic scale compared with the estimated thickness at the same position on the stems.
Figure 8: Illustration of coccolith thickness.
Figure 9

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We thank M.-P. Aubry and J. Henderiks for useful discussions. This work has received financial support from the EU Seventh Framework program Past4Future and from the Agence Nationale de la Recherche under project ANR-12-B06-0007 (CALHIS).

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Authors and Affiliations



L.B. conceived the different aspects of this protocol and wrote the manuscript. N.B. commented on the manuscript, established the equation for the color camera and wrote codes for the color thickness calculation. Y.G. wrote the codes for microscope automation and wrote most of the codes for image analysis. All authors contributed to the protocol and its writing by frequent discussions as well as manuscript reading and correcting.

Corresponding author

Correspondence to Luc Beaufort.

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The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 The Michel-Lévy Interference Color Chart.

This is Figure 2 of Michel-Levy, A. & Lacroix, A. Les Minéraux des roches. 1 : application des méthodes minéralogiques et chimiques à leur étude microscopique. 2 : données physiques et optiques. (Librairie polytechnique Baudry, 1888). It has been found in, a web page written by John Gustav Delly in 2003.

Supplementary information

The Michel-Lévy Interference Color Chart.

This is Figure 2 of Michel-Levy, A. & Lacroix, A. Les Minéraux des roches. 1 : application des méthodes minéralogiques et chimiques ´ leur étude microscopique. 2 : données physiques et optiques. (Librairie polytechnique Baudry, 1888). It has been found in, a web page written by John Gustav Delly in 2003. (PDF 123 kb)

Supplementary Methods

CalciProbe.exe: a macro using the described protocol. (PDF 47 kb)

Supplementary Data

The CalciProbe.exe macro file. (ZIP 712 kb)

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Beaufort, L., Barbarin, N. & Gally, Y. Optical measurements to determine the thickness of calcite crystals and the mass of thin carbonate particles such as coccoliths. Nat Protoc 9, 633–642 (2014).

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