Laser Doppler holographic microscopy in transmission: application to fish embryo imaging Nicolas Verrier, 1 Daniel Alexandre, 1,2 and Michel Gross 1,∗ 1 Laboratoire Charles Coulomb - UMR 5221 CNRS-UM2 CC 026 Universit´ e Montpellier II Place Eug` ene Bataillon 34095 Montpellier cedex, France 2 Montpellier Rio Imaging, Centre de Recherche en Biochimie Macromol´ eculaire - UMR 5237 CNRS-UM1-UM2 1919 route de Mende 34293 Montpellier Cedex 5, France ∗ michel.gross@univ-montp2.fr Abstract: We have extended Laser Doppler holographic microscopy to transmission geometry. The technique is validated with living fish embryos imaged by a modified upright bio-microcope. By varying the frequency of the holographic reference beam, and the combination of frames used to calculate the hologram, multimodal imaging has been performed. Doppler images of the blood vessels for different Doppler shifts, images where the flow direction is coded in RGB colors or movies showing blood cells individual motion have been obtained as well. The ability to select the Fourier space zone that is used to calculate the signal, makes the method quantitative. © 2014 Optical Society of America OCIS codes: (090.1995) Digital holography; (090.2880) Holographic interferometry; (170.3340) Laser Doppler velocimetry; (180.3170) Interference microscopy; (290.5850) Scat- tering, particles; (300.6310) Spectroscopy, heterodyne. References and links 1. E. Friedman, S. Krupsky, A. Lane, S. Oak, E. Friedman, K. Egan, and E. Gragoudas, “Ocular blood flow velocity in age-related macular degeneration,” Ophthalmology 102, 640–646 (1995). 2. M. P. Pase, N. A. Grima, C. K. Stough, A. Scholey, and A. Pipingas, “Cardiovascular disease risk and cerebral blood flow velocity,” Stroke 43, 2803–2805 (2012). 3. J. Kur, E. A. Newman, and T. Chan-Ling, “Cellular and physiological mechanisms underlying blood flow regu- lation in the retina and choroid in health and disease,” Prog. Retin. Eye Res. 31, 377–406 (2012). 4. O. Sakurada, C. Kennedy, J. Jehle, J. Brown, G. L. Carbin, and L. Sokoloff, “Measurement of local cerebral blood flow with iodo [14c] antipyrine,” Am. J. Physiol.-Heart C. 234, H59–H66 (1978). 5. I. Kanno, H. Iida, S. Miura, M. Murakami, K. Takahashi, H. Sasaki, A. Inugami, F. Shishido, and K. Uemura, “A system for cerebral blood flow measurement using an h215o autoradiographic method and positron emission tomography,” J Cerebr. Blood F. Met. 7, 143–153 (1987). 6. Y. Yeh and H. Cummins, “Localized fluid flow measurements with an he-ne laser spectrometer,” Appl. Phys. Lett. 4, 176–178 (1964). 7. J. D. Briers, “Laser doppler, speckle and related techniques for blood perfusion mapping and imaging,” Physiol. Meas. 22, R35–R66 (2001). 8. J. D. Briers and S. Webster, “Laser speckle contrast analysis (lasca): a nonscanning, full-field technique for monitoring capillary blood flow,” J. Biomed. Opt. 1, 174–179 (1996). 9. A. K. Dunn, “Laser speckle contrast imaging of cerebral blood flow,” Ann. Biomed. Eng. 40, 367–377 (2012). 10. D. Briers, D. D. Duncan, E. Hirst, S. J. Kirkpatrick, M. Larsson, W. Steenbergen, T. Stromberg, and O. B. Thompson, “Laser speckle contrast imaging: theoretical and practical limitations,” J. Biomed. Opt. 18, 066018 (2013). 11. S. Yuan, A. Devor, D. A. Boas, and A. K. Dunn, “Determination of optimal exposure time for imaging of blood flow changes with laser speckle contrast imaging,” Appl. Opt. 44, 1823–1830 (2005). #206731 - $15.00 USD Received 19 Feb 2014; revised 28 Mar 2014; accepted 28 Mar 2014; published 10 Apr 2014 (C) 2014 OSA 21 April 2014 | Vol. 22, No. 8 | DOI:10.1364/OE.22.009368 | OPTICS EXPRESS 9368