Subcellular imaging of isotopically labeled carbon compounds in a biological sample by ion microprobe (NanoSIMS)

Abstract

Here we demonstrate the technique of nanoscale secondary ion mass spectrometry, utilizing the Cameca NanoSIMS50 ion microprobe, to detect and image the metabolism of an isotopically labeled compound (NaH(13)CO(3)) in a biological sample. In particular, we have designed and verified protocols for imaging the subcellular distribution and determining the relative abundance of labeled (13)C, within the coral Galaxea fascicularis. Analyses were conducted on 1-mum thick sections of resin-embedded material, using both scanned (mapping) and static (spot analysis) Cs(+) primary ion beam of approximately 100 nm diameter. Using these samples we establish that NanoSIMS has adequate mass resolution to reliably distinguish (13)C from potential isobaric interference by (12)C(1)H and that data extracted from ion maps are comparable to those acquired by spot analyses. Independent of the method of acquisition, ratioing of (13)C to the naturally abundant (12)C is essential if meaningful data, which can be statistically compared to standard and control samples, are to be obtained. These results highlight the potential of NanoSIMS for intracellular tracking of a variety of organic and inorganic compounds labeled with stable isotopes of C, N, O, S, P, and halogens.

Micron-scale mapping of sulfur cycling across the oxycline of a cyanobacterial mat: a paired nanoSIMS and CARD-FISH approach

“…The nanoSIMS has previously been used to analyze sulfur isotopes in minerals (Slodzian et al, 2004;Hoppe et al, 2005) and carbon, nitrogen and sulfur distributions in biological samples (Clode et al, 2007;Li et al, 2008). For the present study, data were obtained using electron multiplier multi-collection, operating with the cesium (Cs þ ) ion source.…”

A single-cell view of ammonium assimilation in coral–dinoflagellate symbiosis

“…In this context, recent advances in a high-resolution isotopic imaging technology, the sub-micrometer scale ion microprobe technique referred to as NanoSIMS provides direct imaging and quantification of the metabolic exchanges within an intact symbiosis following stable isotope labeling (Lechene et al, 2006(Lechene et al, , 2007Kuypers, 2007;Musat et al, 2008;Dattagupta et al, 2009;Foster et al, 2011). Nano-SIMS has recently been successfully applied to reefbuilding corals in order to image trace-elemental distributions in different ultra-structural components of both fossil and living corals (Meibom et al, 2004(Meibom et al, , 2008Clode et al, 2007;Reynaud et al, 2007;Stolarski et al, 2007;Houlbreque et al, 2009;Brahmi et al, 2010). Here, we use NanoSIMS imaging to trace the fixation of isotopically labeled ammonium from seawater within the symbiosis between the reef-building coral Acropora Aspera and the dinoflagellate symbionts Symbiodinium.…”

Electron and ion microprobe analysis of calcium distribution and transport in coral tissues

“…As shown in settled coral larvae by Clode and Marshall (Clode and Marshall, 2004), we show that in mature polyps intracellular total Ca is very high and increases from the outer to inner cell layers. We have also used the ion microprobe at low and high spatial resolution (see Guerquin-Kern et al, 2005;Lechene et al, 2006;Clode et al, 2007) to follow the transport of Ca 2+ across coral epithelia using the stable isotope 44 Ca as…”