Three-dimensional reconstruction of the distribution of elemental tags in single cells using laser ablation ICP-mass spectrometry via registration approaches.
This paper describes a workflow towards the reconstruction of the three-dimensional elemental distribution profile within human cervical carcinoma cells (HeLa), at a spatial resolution down to 1 μm, employing state-of-the-art laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) instrumentation. The suspended cells underwent a series of fixation/embedding protocols and were stained with uranyl acetate and an Ir-based DNA intercalator. A priori, laboratory-based absorption micro-computed tomography (μ-CT) was applied to acquire a reference frame of the morphology of the cells and their spatial distribution before sectioning. After CT analysis, a trimmed 300 × 300 × 300 μm3 block was sectioned into a sequential series of 132 sections with a thickness of 2 μm, which were subjected to LA-ICP-MS imaging. A pixel acquisition rate of 250 pixels s-1 was achieved, through a bidirectional scanning strategy. After acquisition, the two-dimensional elemental images were reconstructed using the timestamps in the laser log file. The synchronization of the data required an improved optimization algorithm, which forces the pixels of scans in different ablation directions to be spatially coherent in the direction orthogonal to the scan direction. The volume was reconstructed using multiple registration approaches. Registration using the section outline itself as a fiducial marker resulted into a volume which was in good agreement with the morphology visualized in the μ-CT volume. The 3D μ-CT volume could be registered to the LA-ICP-MS volume, consisting of 2.9 × 107 voxels, and the nucleus dimensions in 3D space could be derived.
['Van Malderen|Stijn J M|SJM|', 'Van Acker|Thibaut|T|', 'Laforce|Brecht|B|', 'De Bruyne|Michiel|M|', 'de Rycke|Riet|R|', 'Asaoka|Tomoko|T|', 'Vincze|Laszlo|L|', 'Vanhaecke|Frank|F|']
D006367:HeLa Cells D006801:Humans D013058:Mass Spectrometry D059010:Single-Cell Analysis D055114:X-Ray Microtomography
3D HeLa cells Inductively coupled plasma-mass spectrometry (ICP-MS) Laser ablation (LA) Multimodal registration X-ray fluorescence (XRF) spectrometry