Three-dimensional mid-infrared tomographic imaging of endogenous and exogenous molecules in a single intact cell with subcellular resolution.
Microscopy in the mid-infrared spectral range provides detailed chemical information on a sample at moderate spatial resolution and is being used increasingly in the characterization of biological entities as challenging as single cells. However, a conventional cellular 2D imaging measurement is limited in its ability to associate specific compositional information to subcellular structures because of the interference from the complex topography of the sample. Herein we provide a method and protocols that overcome this challenge in which tilt-series infrared tomography is used with a standard benchtop infrared microscope. This approach gives access to the quantitative 3D distribution of molecular components based on the intrinsic contrast provided by the sample. We demonstrate the method by quantifying the distribution of an exogenous metal carbonyl complex throughout the cell and by reporting changes in its coordination sphere in different locations in the cell.
Department of Chemistry, University of Fribourg, Chemin de Musée 9, 1700 Fribourg (Switzerland); Current address: Functional Genomics Center Zurich, Winterthurerstrasse 190, 8057 Zürich (Switzerland). quaroni@bluewin.ch.
Luca
L
Quaroni
Martin
M
Obst
Marcus
M
Nowak
Fabio
F
Zobi
D000490:Allium / Q000737:chemistry / Q000166:cytology* / Q000648:ultrastructure; D021621:Imaging, Three-Dimensional / Q000379:methods*; D007259:Infrared Rays; D008853:Microscopy / Q000379:methods; D059010:Single-Cell Analysis / Q000379:methods*; D013055:Spectrophotometry, Infrared / Q000379:methods*; D041622:Tomography, Optical / Q000379:methods*
chemical imaging; infrared tomography; single-cell studies