title: Vibrational spectroscopic image analysis of biological material using multivariate curve resolution-alternating least squares (MCR-ALS).
abstract: Raman and Fourier transform IR (FTIR) microspectroscopic images of biological material (tissue sections) contain detailed information about their chemical composition. The challenge lies in identifying changes in chemical composition, as well as locating and assigning these changes to different conditions (pathology, anatomy, environmental or genetic factors). Multivariate data analysis techniques are ideal for decrypting such information from the data. This protocol provides a user-friendly pipeline and graphical user interface (GUI) for data pre-processing and unmixing of pixel spectra into their contributing pure components by multivariate curve resolution-alternating least squares (MCR-ALS) analysis. The analysis considers the full spectral profile in order to identify the chemical compounds and to visualize their distribution across the sample to categorize chemically distinct areas. Results are rapidly achieved (usually <30-60 min per image), and they are easy to interpret and evaluate both in terms of chemistry and biology, making the method generally more powerful than principal component analysis (PCA) or heat maps of single-band intensities. In addition, chemical and biological evaluation of the results by means of reference matching and segmentation maps (based on k-means clustering) is possible.
authors.affiliation: Umeå Plant Science Centre, Department of Forest Genetics and Plant Physiology, Swedish University of Agricultural Sciences, Umeå, Sweden.
authors.forename: Judith
authors.initials: J
authors.lastname: Felten
authors.affiliation: Umeå Plant Science Centre, Department of Forest Genetics and Plant Physiology, Swedish University of Agricultural Sciences, Umeå, Sweden.
authors.forename: Hardy
authors.initials: H
authors.lastname: Hall
authors.affiliation: Institut de Diagnòstic Ambiental i Estudis de l'Aigua-Consejo Superior de Investigaciones Científicas (IDAEA-CSIC), Barcelona, Spain.
authors.forename: Joaquim
authors.initials: J
authors.lastname: Jaumot
authors.affiliation: Institut de Diagnòstic Ambiental i Estudis de l'Aigua-Consejo Superior de Investigaciones Científicas (IDAEA-CSIC), Barcelona, Spain.
authors.forename: Romà
authors.initials: R
authors.lastname: Tauler
authors.affiliation: Chemometrics Group, Department of Analytical Chemistry, Universitat de Barcelona, Barcelona, Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: de Juan
authors.affiliation: Department of Chemistry, Umeå University, Umeå, Sweden.
authors.forename: András
authors.initials: A
authors.lastname: Gorzsás
mesh_terms: D000818:Animals; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D007515:Islets of Langerhans / Q000737:chemistry; D016018:Least-Squares Analysis; D008810:Mice, Inbred C57BL; D015999:Multivariate Analysis; D032107:Populus / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared; D013059:Spectrum Analysis, Raman; D014584:User-Computer Interface; D052584:Xylem / Q000737:chemistry