title: Identifying individual cell types in heterogeneous cultures using secondary ion mass spectrometry imaging with C60 etching and multivariate analysis.
abstract: Tissue engineering approaches fabricate and subsequently implant cell-seeded and unseeded scaffold biomaterials. Once in the body, these biomaterials are repopulated with somatic cells of various phenotypes whose identification upon explantation can be expensive and time-consuming. We show that imaging time-of-flight secondary ion mass spectrometry (TOF-SIMS) can be used to distinguish mammalian cell types in heterogeneous cultures. Primary rat esophageal epithelial cells (REEC) were cultured with NIH 3T3 mouse fibroblasts on tissue culture polystyrene and freeze-dried before TOF-SIMS imaging. Results show that a short etching sequence with C(60)(+) ions can be used to clean the sample surface and improve the TOF-SIMS image quality. Principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) were used to identify peaks whose contributions to the total variance in the multivariate model were due to either the two cell types or the substrate. Using PLS-DA, unknown regions of cellularity that were otherwise unidentifiable by SIMS could be classified. From the loadings in the PLS-DA model, peaks were selected that were indicative of the two cell types and TOF-SIMS images were created and overlaid that showed the ability of this method to distinguish features visually.
authors.affiliation: Department of Bioengineering, University of Washington, 3720 15th Ave NE, Box 355061, Seattle, Washington 98195, United States.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Barnes
authors.forename: Jeremy
authors.initials: J
authors.lastname: Brison
authors.forename: Michael
authors.initials: M
authors.lastname: Robinson
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Graham
authors.forename: David G
authors.initials: DG
authors.lastname: Castner
authors.forename: Buddy D
authors.initials: BD
authors.lastname: Ratner
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D003952:Diagnostic Imaging; D004847:Epithelial Cells / Q000378:metabolism*; D004947:Esophagus / Q000166:cytology / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D005612:Freeze Drying; D007091:Image Processing, Computer-Assisted; D016018:Least-Squares Analysis; D051379:Mice; D015999:Multivariate Analysis; D025341:Principal Component Analysis; D051381:Rats; D018629:Spectrometry, Mass, Secondary Ion / Q000379:methods*
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