Approaches to Spectral Imaging Hardware

Jeremy M. Lerner1, Nahum Gat2, Elliot Wachman3

1 LightForm, Inc., Asheville, North Carolina, 2 Opto‐Knowledge Systems, Inc. (OKSI), Torrance, California, 3 ChromoDynamics, Inc., Lakewood, New Jersey
Publication Name:  Current Protocols in Cytometry
Unit Number:  Unit 12.20
DOI:  10.1002/0471142956.cy1220s53
Online Posting Date:  July, 2010
GO TO THE FULL TEXT: PDF or HTML at Wiley Online Library


Instruments used for spectral, multispectral, and hyperspectral imaging in the biosciences have evolved significantly over the last 15 years. However, very few are calibrated and have had their performance validated. Now that spectral imaging systems are appearing in clinics and pathology laboratories, there is a growing need for calibration and validation according to universal standards. In addition, some systems produce spectral artifacts that, at the very least, challenge data integrity if left unrecognized. This unit includes a comparison of the band‐pass and light‐transmission characteristics of electronic tunable filters, interferometers, and wavelength‐dispersive spectral imaging instruments, as well as a description of how they work. Methods are described to test wavelength accuracy and perform radiometric calibration. A real‐life example of spectral artifacts is dissected in detail in order to show how to detect, diagnose, verify, and work around their presence when they cannot be eliminated. Curr. Protoc. Cytom. 53:12.20.1‐12.20.40. © 2009 by John Wiley & Sons, Inc.

Keywords: acousto optic tunable filter; liquid crystal tunable filter; imaging spectrometer; prism spectrometer; diffraction grating ghosts; diffraction grating spectrometer; holographic diffraction grating; ruled diffraction grating

PDF or HTML at Wiley Online Library

Table of Contents

  • Introduction
  • Spectral Imaging Basics
  • Wavelength‐Dispersive Spectral Imaging (WDSI)
  • “Need to Know” Diffraction Grating Characteristics
  • Electronic Tunable Filters (ETF)
  • System‐Based Light Throughput Considerations
  • Wavelength Detectors
  • Calibration, Validation, and Radiometry
  • Comparison of Spectral Imaging Systems
  • Acknowledgements
  • Contact Information
  • Literature Cited
  • Figures
  • Tables
PDF or HTML at Wiley Online Library


PDF or HTML at Wiley Online Library



Literature Cited

Literature Cited
   Cabib, D., Buckwald, R.A., and Nissim, B.‐Y. 1998. Method for chromosome classification by decorrelation statistical analysis and hardware. U.S. Patent No. 5,719,0124.
   Chrisp, M.P. 1987. Aberration‐corrected holographic gratings and their mountings. In Applied Optics and Optical Engineering (R.R. Shannon and W.C. Wyant, eds.) pp. 391‐451. Academic Press, London
   Dicker, D.T., Lerner, J.M., Van Belle, P., Barth, S.F., Guerry, D., Herlyn, M., Elder, D.E. and El‐Deiry, W.S. 2006. Differentiation of normal skin and melanoma using high resolution hyperspectral imaging. Cancer Biol. Ther. 8:1033‐1038.
   Dickinson, M., Bearman, G., Tille, S., Lansford, R., and Fraser, S. 2001. Multi‐spectral imaging and linear unmixing add a whole new dimension to laser scanning fluorescence microscopy. BioTechniques 31:1272‐1278.
   Flamand, J., Bonnemason, F., Lerner, J.M., and Thevenon, A. 1989. Blazing of holographic gratings using ion‐etching. Proc. SPIE 1055:288.
   Garini, Y., Katzir, N., Cabib, D., Buckwald, R., Soenksen, D., and Malik, Z. 1996. Fluorescence imaging spectroscopy and microscopy. In Fluorescence Imaging Spectroscopy and Microscopy (X. Wang and B. Herman, eds.). pp. 87‐124. John Wiley & Sons, New York.
   Gat, N. 2000. Imaging spectroscopy using tunable filters: A review. Proc. SPIE 4056:50‐64.
   Hutley, M.C. 1982. Diffraction Gratings. Academic Press, London.
   James, J. and Sternberg, R. 1969. The Design of Optical Spectrometers. Chapman & Hall, London.
   Lerner, J.M. 2006. Imaging spectrometer fundamentals for researchers in the biosciences: A tutorial. Cytometry A 69:712‐734.
   Lerner, J. and Thevenon, A. 1998. The Optics of Spectroscopy: A Tutorial. Horiba, Metuchen, New Jersey.
   Lerner, J.M. and Zucker, R. 2004. Calibration and validation of confocal spectral imaging systems. Cytometry A 62:8‐34.
   Loewen, E. and Popov, E. 1997. Diffraction Gratings and Applications. Marcel Dekker, New York.
   Miller, P.J. 1991. Use of tunable liquid crystal filters to link radiometric and photometric standards. Metrologia 28:145‐149.
   Prieto‐Blanco, X., Montero‐Orille, C., Couce, B., and de la Fuente, R. 2006. Analytical design of an Offner imaging spectrometer. Opt. Express 14:9156‐9168.
   Reader, J. 1969. Optimizing Czerny‐Turner spectrographs: A comparison between analytical theory and ray tracing. J. Opt. Soc. Am. 59:1189‐1196.
   Shabtay, G., Eidinger, E., Zalevsky, Z., Mendlovic, D., and Marom, E. 2002. Tunable birefringent filters: Optimal iterative design. Opt. Express 10:1534‐1541.
   Stroke, G.W. 1967. Diffraction gratings (in English). In Handbuch der Physik XXIX (S. Flügge, ed.), vol. 29. Springer‐Verlag, Berlin.
   Tumeh, P.C., Lerner, J.M., Dicker, D.T., and El‐Deiry, W.S. 2007. Differentiation of vascular and non‐vascular skin spectral signatures using in vivo hyperspectral radiometric imaging: Implications for monitoring angiogenesis. Cancer Biol. Ther. 6:447‐453.
   van der Loos, C.M. 2008. Multiple immunoenzyme staining: Methods and visualizations for the observation with spectral imaging. J. Immunochem. Cytochem. 56:313‐328.
   Vo‐Dinh, T. 2004. A hyperspectral imaging system for in vivo optical diagnostics. IEEE Engin. Med. Biol. Mag. 23:40‐49.
   Wachman, E.S. and Pannell, C.N. 2008. High‐performance hyperspectral imager using a novel acousto‐optic tuneable filter. Proc. SPIE 6966:7‐12.
   Warren, D., Hackwell, J., and Gutierrez, D. 1997. Compact prism spectrographs based on aplanatic principles. Opt. Eng. 36:1174‐1182.
   Ye, C. 2003. Wavelength‐tunable spectral filters based on the optical rotatory dispersion effect. Appl. Optics 42:4505‐4513.
PDF or HTML at Wiley Online Library