title: Automated cell-by-cell tissue imaging and single-cell analysis for targeted morphologies by laser ablation electrospray ionization mass spectrometry.
abstract: Mass spectrometry imaging (MSI) is an emerging technology for the mapping of molecular distributions in tissues. In most of the existing studies, imaging is performed by sampling on a predefined rectangular grid that does not reflect the natural cellular pattern of the tissue. Delivering laser pulses by a sharpened optical fiber in laser ablation electrospray ionization (LAESI) mass spectrometry (MS) has enabled the direct analysis of single cells and subcellular compartments. Cell-by-cell imaging had been demonstrated using LAESI-MS, where individual cells were manually selected to serve as natural pixels for tissue imaging. Here we describe a protocol for a novel cell-by-cell LAESI imaging approach that automates cell recognition and addressing for systematic ablation of individual cells. Cell types with particular morphologies can also be selected for analysis. First, the cells are recognized as objects in a microscope image. The coordinates of their centroids are used by a stage-control program to sequentially position the cells under the optical fiber tip for laser ablation. This approach increases the image acquisition efficiency and stability, and enables the investigation of extended or selected tissue areas. In the LAESI process, the ablation events result in mass spectra that represent the metabolite levels in the ablated cells. Peak intensities of selected ions are used to represent the metabolite distributions in the tissue with single-cell resolution.
authors.affiliation: Department of Chemistry, W. M. Keck Institute for Proteomics Technology and Applications, The George Washington University, 725 21-st Street, N.W., Washington, DC, 20052, USA.
authors.forename: Hang
authors.initials: H
authors.lastname: Li
authors.forename: Brian K
authors.initials: BK
authors.lastname: Smith
authors.forename: Bindesh
authors.initials: B
authors.lastname: Shrestha
authors.forename: László
authors.initials: L
authors.lastname: Márk
authors.forename: Akos
authors.initials: A
authors.lastname: Vertes
mesh_terms: D000490:Allium / Q000166:cytology; D001331:Automation; D053685:Laser Therapy / Q000295:instrumentation / Q000379:methods*; D027762:Lilium / Q000166:cytology; D057054:Molecular Imaging / Q000379:methods*; D055100:Optical Fibers; D059010:Single-Cell Analysis / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*