title: Three-dimensional mass spectral imaging of HeLa-M cells--sample preparation, data interpretation and visualisation.
abstract: Time-of-flight secondary ion mass spectrometry (ToFSIMS) is being applied increasingly to the study of biological systems where the chemical specificity of mass spectrometry and the high lateral resolution imaging capabilities can be exploited. Here we report a comparison of two cell sample preparation methods and demonstrate how they influence the outcome of the ToFSIMS analysis for three-dimensional (3D) imaging of biological cells using our novel buncher-ToF instrument (J105 3D Chemical Imager) equipped with a C(60) primary ion beam. Cells were analysed fixed and freeze-dried and non-fixed, frozen-hydrated. It is concluded that maintaining the cells in a non-fixed frozen-hydrated state during the analysis helps reduce chemical redistribution, producing cleaner spectra and improved chemical contrast in both 2D and 3D imaging. Insights into data interpretation are included and we present methods for 3D reconstruction of the data using multivariate analysis techniques.
authors.affiliation: Manchester Interdisciplinary Biocentre, School of Chemical Engineering and Analytical Science, University of Manchester, 131 Princess Street, Manchester M1 7DN, UK. john.fletcher@manchester.ac.uk
authors.forename: John S
authors.initials: JS
authors.lastname: Fletcher
authors.forename: Sadia
authors.initials: S
authors.lastname: Rabbani
authors.forename: Alex
authors.initials: A
authors.lastname: Henderson
authors.forename: Nicholas P
authors.initials: NP
authors.lastname: Lockyer
authors.forename: John C
authors.initials: JC
authors.lastname: Vickerman
mesh_terms: D000225:Adenine / Q000737:chemistry; D003584:Cytological Techniques / Q000295:instrumentation / Q000379:methods*; D005614:Freeze Fracturing; D006367:HeLa Cells; D006801:Humans; D021621:Imaging, Three-Dimensional / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D057054:Molecular Imaging / Q000379:methods*; D015999:Multivariate Analysis; D010767:Phosphorylcholine / Q000737:chemistry; D025341:Principal Component Analysis
chemical_list: D010767:Phosphorylcholine; D000225:Adenine