title: Noninvasive detection and imaging of molecular markers in live cardiomyocytes derived from human embryonic stem cells.
abstract: Raman microspectroscopy (RMS) was used to detect and image molecular markers specific to cardiomyocytes (CMs) derived from human embryonic stem cells (hESCs). This technique is noninvasive and thus can be used to discriminate individual live CMs within highly heterogeneous cell populations. Principal component analysis (PCA) of the Raman spectra was used to build a classification model for identification of individual CMs. Retrospective immunostaining imaging was used as the gold standard for phenotypic identification of each cell. We were able to discriminate CMs from other phenotypes with >97% specificity and >96% sensitivity, as calculated with the use of cross-validation algorithms (target 100% specificity). A comparison between Raman spectral images corresponding to selected Raman bands identified by the PCA model and immunostaining of the same cells allowed assignment of the Raman spectral markers. We conclude that glycogen is responsible for the discrimination of CMs, whereas myofibril proteins have a lesser contribution. This study demonstrates the potential of RMS for allowing the noninvasive phenotypic identification of hESC progeny. With further development, such label-free optical techniques may enable the separation of high-purity cell populations with mature phenotypes, and provide repeated measurements to monitor time-dependent molecular changes in live hESCs during differentiation in vitro.
authors.affiliation: School of Physics and Astronomy, University of Nottingham, Nottingham, UK.
authors.forename: Flavius C
authors.initials: FC
authors.lastname: Pascut
authors.forename: Huey T
authors.initials: HT
authors.lastname: Goh
authors.forename: Nathan
authors.initials: N
authors.lastname: Welch
authors.forename: Lee D
authors.initials: LD
authors.lastname: Buttery
authors.forename: Chris
authors.initials: C
authors.lastname: Denning
authors.forename: Ioan
authors.initials: I
authors.lastname: Notingher
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002470:Cell Survival; D053595:Embryonic Stem Cells / Q000166:cytology*; D006801:Humans; D007834:Lasers; D051379:Mice; D057054:Molecular Imaging / Q000379:methods*; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000378:metabolism*; D010641:Phenotype; D025341:Principal Component Analysis; D013059:Spectrum Analysis, Raman
chemical_list: D015415:Biomarkers