An improved energy-dispersive X-ray microanalysis method for analyzing simultaneously carbon, nitrogen, oxygen, phosphorus, sulfur, and other cation and anion concentrations in single natural marine microplankton cells

Mariona Segura-Noguera, Dolors Blasco, José-Manuel Fortuño

Limnol. Oceanogr. Methods 10:666-680 (2012) | DOI: 10.4319/lom.2012.10.666

ABSTRACT: Electron-probe X-ray microanalysis (XRMA) is a technique that can be used to simultaneously quantify C, N, O, Na, Mg, Al, Si, P, S, Cl, K, and Ca present in single cells collected from the sea, with a minimal treatment of the samples. The technique has been used to determine elemental composition in plankton using scanning electron microscopes (SEM) or transmission electron microscopes (TEM). The methodologies used have shown some drawbacks. Here we present an improved methodology focusing on the analysis of the elemental concentration of marine microplankton cells, including light elements, based on the methodology of Norland et al. (1995). The most important modification is the use of a SEM microscope but reproducing TEM conditions by placing the sample grid at a distance from the holder bottom. This modification eliminates the interference of the SEM stub from the analysis of the cells, and improves the limit of detection of elements that are present in low concentrations. Experimental and theoretical data on the beam penetration depth using low incident beam energies (15 kV) are presented. The results show that a 15 kV accelerating voltage, invoked as a limitation in using XRMA for the analysis of samples thicker than 0.1 μm (Twining et al. 2008), is sufficient to analyze whole microplankton cells, and that there is no absorption of X-rays during the analysis. The method has been applied to diatoms and dinoflagellates from the Western Mediterranean Sea. The cell elemental composition results fall within the range of historical data in the literature.