title: Recognizing different tissues in human fetal femur cartilage by label-free Raman microspectroscopy.
abstract: Traditionally, the composition of bone and cartilage is determined by standard histological methods. We used Raman microscopy, which provides a molecular "fingerprint" of the investigated sample, to detect differences between the zones in human fetal femur cartilage without the need for additional staining or labeling. Raman area scans were made from the (pre)articular cartilage, resting, proliferative, and hypertrophic zones of growth plate and endochondral bone within human fetal femora. Multivariate data analysis was performed on Raman spectral datasets to construct cluster images with corresponding cluster averages. Cluster analysis resulted in detection of individual chondrocyte spectra that could be separated from cartilage extracellular matrix (ECM) spectra and was verified by comparing cluster images with intensity-based Raman images for the deoxyribonucleic acid/ribonucleic acid (DNA/RNA) band. Specific dendrograms were created using Ward's clustering method, and principal component analysis (PCA) was performed with the separated and averaged Raman spectra of cells and ECM of all measured zones. Overall (dis)similarities between measured zones were effectively visualized on the dendrograms and main spectral differences were revealed by PCA allowing for label-free detection of individual cartilaginous zones and for label-free evaluation of proper cartilaginous matrix formation for future tissue engineering and clinical purposes.
authors.affiliation: University of Twente, MIRA-Institute for Biomedical Technology and Technical Medicine, Department of Tissue Regeneration, Drienerlolaan 5, 7522 NB Enschede, The Netherlands.
authors.forename: Aliz
authors.initials: A
authors.lastname: Kunstar
authors.forename: Jeroen
authors.initials: J
authors.lastname: Leijten
authors.forename: Stefan
authors.initials: S
authors.lastname: van Leuveren
authors.forename: Janneke
authors.initials: J
authors.lastname: Hilderink
authors.forename: Cees
authors.initials: C
authors.lastname: Otto
authors.forename: Clemens A
authors.initials: CA
authors.lastname: van Blitterswijk
authors.forename: Marcel
authors.initials: M
authors.lastname: Karperien
authors.forename: Aart A
authors.initials: AA
authors.lastname: van Apeldoorn
mesh_terms: D002358:Cartilage, Articular / Q000033:anatomy & histology* / Q000737:chemistry; D019902:Chondrocytes / Q000737:chemistry / Q000166:cytology; D005109:Extracellular Matrix / Q000737:chemistry / Q000648:ultrastructure; D005269:Femur / Q000033:anatomy & histology* / Q000737:chemistry*; D005333:Fetus / Q000033:anatomy & histology* / Q000737:chemistry*; D006132:Growth Plate / Q000033:anatomy & histology / Q000737:chemistry; D006801:Humans; D055620:Optical Phenomena; D025341:Principal Component Analysis; D013059:Spectrum Analysis, Raman / Q000379:methods*