title: Non-invasive label-free monitoring the cardiac differentiation of human embryonic stem cells in-vitro by Raman spectroscopy.
abstract: BACKGROUND
  Online label-free monitoring of in-vitro differentiation of stem cells remains a major challenge in stem cell research. In this paper we report the use of Raman micro-spectroscopy (RMS) to measure time- and spatially-resolved molecular changes in intact embryoid bodies (EBs) during in-vitro cardiogenic differentiation.
  
  
  METHODS
  EBs formed by aggregation of human embryonic stem cells (hESCs) were cultured in defined medium to induce differentiation towards cardiac phenotype and maintained in purpose-built micro-bioreactors on the Raman microscope for 5days (between days 5 and 9 of differentiation) and spatially-resolved spectra were recorded at 24h intervals.
  
  
  RESULTS
  The Raman spectra showed that the onset of spontaneous beating of EBs at day 7 coincided with an increase in the intensity of the Raman bands at 1340cm(-1), 1083cm(-1), 937cm(-1), 858cm(-1), 577cm(-1) and 482cm(-1). The spectral maps corresponding to these bands had a high positive correlation with the expression of the cardiac-specific α-actinin obtained by immuno-fluorescence imaging of the same EBs. The spectral markers obtained here are also in agreement with previous studies performed on individual live hESC-derived CMs.
  
  
  CONCLUSIONS
  The intensity profile of these Raman bands can be used for label-free in-situ monitoring of EBs to estimate the efficacy of cardiogenic differentiation.
  
  
  CONCLUSIONS
  As the acquisition of the time-course Raman spectra did not affect the viability or the differentiation potential of the hESCs, this study demonstrates the feasibility of using RMS for on-line non-invasive continuous monitoring of such processes inside bioreactor culture systems.
authors: School of Physics and Astronomy, University of Nottingham, Nottingham, NG7 2RD, UK.
authors: Flavius C
authors: FC
authors: Pascut
authors: Spandan
authors: S
authors: Kalra
authors: Vinoj
authors: V
authors: George
authors: Nathan
authors: N
authors: Welch
authors: Chris
authors: C
authors: Denning
authors: Ioan
authors: I
authors: Notingher
mesh_terms: D000185:Actinin / Q000096:biosynthesis; D000943:Antigens, Differentiation / Q000096:biosynthesis; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D006801:Humans; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism; D013059:Spectrum Analysis, Raman / Q000379:methods*
chemical_list: D000943:Antigens, Differentiation; D000185:Actinin