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

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.

Taxonomic and Environmental Variability in the Elemental Composition and Stoichiometry of Individual Dinoflagellate and Diatom Cells from the NW Mediterranean Sea

“…Unfortunately, data on the elemental composition of plankton in nature is still too sparse to validate these hypotheses [ 9 , 12 ], specially for particulate phosphorus [ 13 ]. Two relatively new methods, energy dispersive X-ray microanalysis (XRMA, also abbreviated as EDX or EDS for energy dispersive X-ray spectroscopy) [ 14 , 15 ] and synchrotron–based X-ray fluorescence microprobe (SXRF) [ 16 ], show promise to overcome this scarcity. However, these methods are not yet routinely applied and the existing data is still limited, available only for a few taxonomic groups and environmental conditions.…”

X ray microanalysis of northeastern Quintana Roo aquatic biota, Mexico: evidence of hazard metals presence

“…In this way, the biota and its elemental composition are related to the hydrogeochemistry of water. For example, in diatoms and marine dinoflagellates, the presence of Ca, K, Mg, P, S and Si has been reported as part of the composition of their body structure (Segura-Noguera et al, 2012). Also, variations in the concentrations of Ca, Cl, K, Na, Mg, P, S and Si in Ceratium hirundinella are caused by seasonality (Sigee et al, 1998).…”

“…The X-ray penetration range of 1,000 nm. The average penetration range in X-ray analyses is 2.275 µm (Segura-Noguera et al, 2012). INCA suite 3.04 was used for the quantitative analyses (the total composition percent of the detected elements).…”

Evaluating the stoichiometric trait distributions of cultured bacterial populations and uncultured microbial communities

“…However, these studies commonly reported averaged trait values and did not present the distribution of these stoichiometric traits. Since these early studies, the equipment and techniques used for EDS have improved and enabled single-cell stoichiometries to be detected and reported with greater sensitivity and in higher resolution (Segura-Noguera et al, 2012;Watanabe, 2013). Here, we aim to advance the description of the stoichiometric trait distribution for cultured and uncultured bacteria by integrating single cell EDS-generated stoichiometric data into probability density functions to describe the frequency distribution of stoichiometric traits of individual bacterial cells within a population or community.…”