Skip to main content
Log in

Isolating stem cells in the inter-follicular epidermis employing synchrotron radiation-based Fourier-transform infrared microspectroscopy and focal plane array imaging

  • Original Paper
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript


Normal function and physiology of the epidermis is maintained by the regenerative capacity of this tissue via adult stem cells (SCs). However, definitive identifying markers for SCs remain elusive. Infrared (IR) spectroscopy exploits the ability of cellular biomolecules to absorb in the mid-IR region (λ = 2.5–25 μm), detecting vibrational transitions of chemical bonds. In this study, we exploited the cell’s inherent biochemical composition to discriminate SCs of the inter-follicular skin epidermis based on IR-derived markers. Paraffin-embedded samples of human scalp skin (n = 4) were obtained, and 10-μm thick sections were mounted for IR spectroscopy. Samples were interrogated in transmission mode using synchrotron radiation-based Fourier-transform IR (FTIR) microspectroscopy (15 × 15 μm) and also imaged employing globar-source FTIR focal plane array (FPA) imaging (5.4 × 5.4 μm). Dependent on the location of derived spectra, wavenumber–absorbance/intensity relationships were examined using unsupervised principal component analysis. This approach showed clear separation and spectral differences dependent on cell type. Spectral biomarkers concurrently associated with segregation of SCs, transit-amplifying cells and terminally-differentiated cells of epidermis were primarily PO 2 vibrational modes (1,225 and 1,080 cm−1), related to DNA conformational alterations. FPA imaging coupled with hierarchical cluster analysis also indicated the presence of specific basal layer cells potentially originating from the follicular bulge, suggested by co-clustering of spectra. This study highlights PO 2 vibrational modes as potential putative SC markers.

“Delineating the putative stem cell lineage in interfollicular skin based on position-derived infrared spectral fingerprints”.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others



Focal plane array


Fourier-transform infrared


Haematoxylin and eosin


Hierarchical cluster analysis




Linear discriminant analysis


Numerical aperture


Principal component


Principal component analysis


stem cell


Signal-to-noise ratio





νasPO 2 :

Asymmetric phosphate stretching vibrations

νsPO 2 :

Symmetric phosphate stretching vibrations


  1. Patel II, Trevisan J, Singh PB, Nicholson CM, Krishnan RK, Matanhelia SS, Martin FL (2011) Segregation of human prostate tissues classified high-risk (UK) versus low-risk (India) for adenocarcinoma using Fourier-transform infrared or Raman microspectroscopy coupled with discriminant analysis. Anal Bioanal Chem 401:969–982

    Article  CAS  Google Scholar 

  2. Martin FL, Kelly JG, Llabjani V, Martin-Hirsch PL, Patel II, Trevisan J, Fullwood NJ, Walsh MJ (2010) Distinguishing cell types or populations based on the computational analysis of their infrared spectra. Nat Protoc 5:1748–1760

    Article  CAS  Google Scholar 

  3. Kelly JG, Angelov PP, Trevisan J, Vlachopoulou A, Paraskevaidis E, Martin-Hirsch PL, Martin FL (2010) Robust classification of low-grade cervical cytology following analysis with ATR-FTIR spectroscopy and subsequent application of self-learning classifier eClass. Anal Bioanal Chem 398:2191–2201

    Article  CAS  Google Scholar 

  4. Patel II, Trevisan J, Evans G, Llabjani V, Martin Hirsh PL, Stringfellow HF, Martin FL (2011) High contrast images of uterine tissue derived using Raman microspectroscopy with the empty modelling approach of multivariate curve resolution-alternating least squares. Analyst 136:4950–4959

    Article  CAS  Google Scholar 

  5. Fuchs E (2008) Skin stem cells: rising to the surface. J Cell Biol 180:273–284

    Article  CAS  Google Scholar 

  6. Li L, Clevers H (2010) Coexistence of quiescent and active adult stem cells in mammals. Science 29:542–545

    Article  Google Scholar 

  7. Menon GK (2002) New insights into skin structure: scratching the surface. Adv Drug Deliv Rev 54:S3–S17

    Article  CAS  Google Scholar 

  8. Morasso MI, Tomic-Canic M (2005) Epidermal stem cells: the cradle of epidermal determination, differentiation and wound healing. Biol Cell 97:173–183

    Article  CAS  Google Scholar 

  9. Fuchs E, Horsley V (2008) More than one way to skin. Genes Dev 22:976–985

    Article  CAS  Google Scholar 

  10. Pajoohesh-Ganji A, Stepp MA (2005) In search of markers for the stem cells of the corneal epithelium. Biol Cell 97:265–276

    Article  CAS  Google Scholar 

  11. Pincelli C, Marconi A (2010) Keratinocyte stem cells: friends and foes. J Cell Physiol 225:310–315

    Article  CAS  Google Scholar 

  12. Voog J, Jones DL (2010) Stem cells and the niche: a dynamic duo. Cell Stem Cell 6:103–115

    Article  CAS  Google Scholar 

  13. Stenn KS, Paus R (2001) Controls of hair follicle cycling. Physiol Rev 81:449–494

    CAS  Google Scholar 

  14. Lai-Cheong JE, McGrath JA (2009) Structure and function of skin, hair and nails. Medicine 37:223–226

    Article  Google Scholar 

  15. Blanpain C, Lowry WE, Geoghegan A, Polak L, Fuchs E (2004) Self-renewal, multipotency, and the existence of two cell populations within an epithelial stem cell niche. Cell 118:635–648

    Article  CAS  Google Scholar 

  16. Jaks V, Barker N, Kasper M, van Es JH, Snippert HJ, Clevers H, Toftgård R (2008) Lgr5 marks cycling, yet long-lived, hair follicle stem cells. Nat Genet 40:1291–1299

    Article  CAS  Google Scholar 

  17. Jaks V, Kasper M, Toftgård R (2010) The hair follicle-a stem cell zoo. Exp Cell Res 316:1422–1428

    Article  CAS  Google Scholar 

  18. Walsh MJ, Fellous TG, Hammiche A, Lin WR, Fullwood NJ, Grude O, Bahrami F, Nicholson JM, Cotte M, Susini J, Pollock HM, Brittan M, Martin-Hirsch PL, Alison MR, Martin FL (2008) Fourier transform infrared microspectroscopy identifies symmetric PO 2 modifications as a marker of the putative stem cell region of human intestinal crypts. Stem Cells 26:108–118

    Article  CAS  Google Scholar 

  19. Walsh MJ, Hammiche A, Fellous TG, Nicholson JM, Cotte M, Susini J, Fullwood NJ, Martin-Hirsch PL, Alison MR, Martin FL (2009) Tracking the cell hierarchy in the human intestine using biochemical signatures derived by mid-infrared microspectroscopy. Stem Cell Res 3:15–27

    Article  CAS  Google Scholar 

  20. Chan JW, Lieu DK (2009) Label-free biochemical characterization of stem cells using vibrational spectroscopy. J Biophotonics 2:656–668

    Article  CAS  Google Scholar 

  21. Bentley AJ, Nakamura T, Hammiche A, Martin FL, Pollock HM, Kinoshito S, Fullwood NJ (2009) Characterization of human corneal stem cells by synchrotron infrared micro-spectroscopy. Mol Vis 13:237–242

    Google Scholar 

  22. Grude O, Nakamura T, Hammiche A, Bentley AJ, Martin FL, Pollock HM, Kinoshito S, Fullwood NJ (2009) Discrimination of human stem cells by photothermal microspectroscopy. Vib Spectrosc 49:22–27

    Article  CAS  Google Scholar 

  23. Kelly JG, Nakamura T, Kinoshita S, Fullwood NJ, Martin FL (2010) Evidence for a stem-cell lineage in corneal squamous cell carcinoma using synchrotron-based Fourier-transform infrared microspectroscopy and multivariate analysis. Analyst 135:3120–3125

    Article  CAS  Google Scholar 

  24. Nasse MJ, Walsh MJ, Mattson EC, Reininger R, Kajdacsy-Balla A, Macias V, Bhargava R, Hirschmugl CJ (2011) High-resolution Fourier-transform infrared chemical imaging with multiple synchrotron beams. Nat Methods 8:413–416

    Article  CAS  Google Scholar 

  25. Thumanu K, Tanthanuch W, Lorthongpanich C, Heraud P, Parnpai R (2009) FTIR microspectroscopic imaging as a new tool to distinguish chemical composition of mouse blastocyst. J Mol Struct 933:104–111

    Article  CAS  Google Scholar 

  26. Heraud P, Caine S, Campanale N, Karnezis T, McNaughton D, Wood BR, Tobin MJ, Bernard CC (2010) Early detection of the chemical changes occurring during the induction and prevention of autoimmune-mediated demyelination detected by FT-IR imaging. NeuroImage 49:1180–1189

    Article  Google Scholar 

  27. Hobro AJ, Lendl B (2011) Fourier-transform mid-infrared FPA imaging of a complex multicellular nematode. Vib Spectrosc 57:213–219

    Article  CAS  Google Scholar 

  28. Lau K, Hedegaard MA, Kloepper JE, Paus R, Wood BR, Deckert V (2011) Visualization and characterisation of defined hair follicle compartments by Fourier transform infrared (FTIR) imaging without labelling. J Dermatol Sci 63:191–198

    Article  CAS  Google Scholar 

  29. Bambery KR, Wood BR, McNaughton D (2012) Resonant Mie scattering (RMieS) correction applied to FTIR images of biological tissue samples. Analyst 137:126–132

    Article  CAS  Google Scholar 

  30. Harrison WJ, Bull JJ, Seltmann H, Zouboulis CC, Philpott MP (2007) Expression of lipogenic factors galectin-12, resistin, SREBP-1, and SCD in human sebaceous glands and cultured sebocytes. J Investig Dermatol 127:1309–1317

    Article  CAS  Google Scholar 

  31. Bassan P, Byrne HJ, Bonnier F, Lee J, Dumas P, Gardner P (2009) Resonant Mie scattering in infrared spectroscopy of biological materials—understanding the ‘dispersion artefact’. Analyst 134:1586–1593

    Article  CAS  Google Scholar 

  32. Bassan P, Kohler A, Martens H, Lee J, Jackson E, Lockyer N, Dumas P, Brown M, Clarke N, Gardner P (2010) RMieS-EMSC correction for infrared spectra of biological cells: extension using full Mie theory and GPU computing. J Biophotonics 3:609–620

    Article  CAS  Google Scholar 

  33. Martin FL, German MJ, Wit E, Fearn T, Ragavan N, Pollock HM (2007) Identifying variables responsible for clustering in discriminant analysis of data from infrared microspectroscopy of a biological sample. J Comput Biol 14:1176–1184

    Article  CAS  Google Scholar 

  34. Watt FM, Fujiwara H (2011) Cell-extracellular matrix interactions in normal and diseased skin. Cold Spring Harb Perspect Biol 3. doi:10.1101/cshperspect.a005124

  35. Nakamura T, Kelly JG, Trevisan J, Cooper LJ, Bentley AJ, Carmichael PL, Scott AD, Cotte M, Susini J, Martin-Hirsch PL, Kinoshita S, Fullwood NJ, Martin FL (2010) Microspectroscopy of spectral biomarkers associated with human corneal stem cells. Mol Vis 16:359–368

    CAS  Google Scholar 

  36. Moll R, Divo M, Langbein L (2008) The human keratins: biology and pathology. Histochem Cell Biol 129:705–733

    Article  CAS  Google Scholar 

  37. Kelly JG, Trevisan J, Scott AD, Carmichael PL, Pollock HM, Martin-Hirsch PL, Martin FL (2011) Biospectroscopy to metabolically profile biomolecular structure: a multistage approach linking computational analysis with biomarkers. J Proteome Res 10:1437–1448

    Article  CAS  Google Scholar 

  38. Trevisan J, Angelov PP, Carmichael PL, Scott AD, Martin FL (2012) Extracting biological information with computational analysis of Fourier-transform infrared (FTIR) biospectroscopy datasets: current practices to future perspectives. Analyst 137:3202–3215

    Article  CAS  Google Scholar 

Download references


This work was funded by Rosemere Cancer Foundation and Unilever PLC. We also thanks the Sciences and Technologies Facilities Council for grant support to access the Diamond synchrotron facility.

Disclosure of potential conflicts of interest

The authors indicate no potential conflicts of interest.

Author information

Authors and Affiliations


Corresponding author

Correspondence to Francis L. Martin.

Electronic supplementary material

Below is the link to the electronic supplementary material.


(PDF 412 KB)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Patel, I.I., Harrison, W.J., Kerns, J.G. et al. Isolating stem cells in the inter-follicular epidermis employing synchrotron radiation-based Fourier-transform infrared microspectroscopy and focal plane array imaging. Anal Bioanal Chem 404, 1745–1758 (2012).

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: