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Compatibility of Staining Protocols for Bone Tissue with Raman Imaging

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Abstract

We report the use of Raman microscopy to image mouse calvaria stained with hematoxylin, eosin and toluidine blue. Raman imaging of stained specimens allows for direct correlation of histological and spectral information. A line-focus 785 nm laser imaging system with specialized near-infrared (NIR) microscope objectives and CCD detector were used to collect approximately 100 × 450 µm Raman images. Principal components analysis, a multivariate analysis technique, was used to determine whether the histological stains cause spectral interference (band shifts or intensity changes) or result in thermal damage to the examined tissue. Image analysis revealed factors for tissue components and the embedding medium, glycol methacrylate, only. Thus, Raman imaging proved to be compatible with histological stains such as hematoxylin, eosin and toluidine blue.

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Acknowledgements

Research was supported by NIH DE11530 (to M.A.I), NIH AR47969 (to M.D.M.), U.S. Army Research Office DAMD17-01-1-0809 (to M.D.M.), and University of Michigan Rackham School of Graduate Studies Summer Research Opportunity Program Fellowship (L.E.G.). The authors would like to thank John Baker (University of Michigan) for his assistance with sample preparation.

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Authors and Affiliations

  1. Department of Chemistry, University of Michigan, 930 N. University Avenue, Ann Arbor, MI 48109-1055, USA

    Michael D. Morris & Nicole J. Crane

  2. Department of Orthodontics and Pediatric Dentistry, University of Michigan School of Dentistry, Ann Arbor, MI, USA

    Luis E. Gomez & Michael A. Ignelzi Jr

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  1. Michael D. Morris
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  2. Nicole J. Crane
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  3. Luis E. Gomez
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  4. Michael A. Ignelzi Jr
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Correspondence to Michael D. Morris.

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Morris, M.D., Crane, N.J., Gomez, L.E. et al. Compatibility of Staining Protocols for Bone Tissue with Raman Imaging . Calcif Tissue Int 74, 86–94 (2004). https://doi.org/10.1007/s00223-003-0038-0

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  • DOI: https://doi.org/10.1007/s00223-003-0038-0

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