Identifying Individual Cell Types in Heterogeneous Cultures Using SIMS Imaging with C60 Etching and Multivariate Analysis

摘要

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 C60⁺ 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 of 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.