Hostname: page-component-76fb5796d-9pm4c Total loading time: 0 Render date: 2024-04-29T18:13:42.924Z Has data issue: false hasContentIssue false
  • English
  • Français

Two-Color, Two-Photon Imaging at Long Excitation Wavelengths Using a Diamond Raman Laser

Published online by Cambridge University Press:  05 August 2016

Johanna Trägårdh Open the ORCID record for Johanna Trägårdh [Opens in a new window] ,
Michelle Murtagh ,
Gillian Robb ,
Maddy Parsons ,
Jipeng Lin ,
David J. Spence  and
Gail McConnell
Johanna Trägårdh*
Affiliation:
Centre for Biophotonics, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 161 Cathedral Street, Glasgow, G4 0RE, UK
Michelle Murtagh
Affiliation:
Centre for Biophotonics, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 161 Cathedral Street, Glasgow, G4 0RE, UK MQ Photonics, Department of Physics and Astronomy, Macquarie University, NSW 2109, Australia
Gillian Robb
Affiliation:
Centre for Biophotonics, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 161 Cathedral Street, Glasgow, G4 0RE, UK
Maddy Parsons
Affiliation:
Randall Division of Cell and Molecular Biophysics, King’s College London, Guy’s Campus, London, SE11UL, UK
Jipeng Lin
Affiliation:
MQ Photonics, Department of Physics and Astronomy, Macquarie University, NSW 2109, Australia
David J. Spence
Affiliation:
MQ Photonics, Department of Physics and Astronomy, Macquarie University, NSW 2109, Australia
Gail McConnell
Affiliation:
Centre for Biophotonics, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 161 Cathedral Street, Glasgow, G4 0RE, UK
*
*Corresponding author. johanna.tragardh@strath.ac.uk
Get access

Abstract

We demonstrate that the second-Stokes output from a diamond Raman laser, pumped by a femtosecond Ti:Sapphire laser, can be used to efficiently excite red-emitting dyes by two-photon excitation at 1,080 nm and beyond. We image HeLa cells expressing red fluorescent protein, as well as dyes such as Texas Red and Mitotracker Red. We demonstrate the potential for simultaneous two-color, two-photon imaging with this laser by using the residual pump beam for excitation of a green-emitting dye. We demonstrate this for the combination of Alexa Fluor 488 and Alexa Fluor 568. Because the Raman laser extends the wavelength range of the Ti:Sapphire laser, resulting in a laser system tunable to 680–1,200 nm, it can be used for two-photon excitation of a large variety and combination of dyes.

Keywords

nonlinear microscopyfluorescence microscopyRaman lasersfluorescent proteinstwo-photon microscopy
Type
Technique and Instrumentation Development
Information
Microscopy and Microanalysis , Volume 22 , Issue 4 , August 2016 , pp. 803 - 807
Copyright
© Microscopy Society of America 2016 

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Anderson, R.R. & Parrish, J.A. (1981). The optics of human skin. J Invest Dermatol 77, 13–19.CrossRefGoogle ScholarPubMed
Bradley, D.J., Hutchinson, M.H.R., Koetser, H., Morrow, G., New, G.H.C. & Petty, M.S. (1972). Interactions of picosecond laser pulses with organic molecules. I. two-photon fluorescence quenching and singlet states excitation in rhodamine dyes. Proc R Soc Lond A 328, 97–121.Google Scholar
Brenner, M.H., Cai, D., Swanson, J.A. & Ogilvie, J.P. (2013). Two-photon imaging of multiple fluorescent proteins by phase-shaping and linear unmixing with a single broadband laser. Opt Express 21, 17256–17264.CrossRefGoogle ScholarPubMed
Butko, M.T., Drobizhev, M., Makarov, N.S., Rebane, A., Brinkman, B.C. & Gleeson, J.G. (2011). Simultaneous multiple-excitation multiphoton microscopy yields increased imaging sensitivity and specificity. BMC Biotechnol 11, 20.CrossRefGoogle ScholarPubMed
Chu, S.W., Chen, I.H., Liu, T.M., Chen, P.C., Sun, C.K. & Lin, B.L. (2001). Multimodal nonlinear spectral microscopy based on a femtosecond Cr:forsterite laser. Opt Lett 26, 1909–1911.CrossRefGoogle ScholarPubMed
Churin, D., Olson, J., Norwood, R.A., Peyghambarian, N. & Kieu, K. (2015). High-power synchronously pumped femtosecond Raman fiber laser. Opt Lett 40, 2529–2532.CrossRefGoogle ScholarPubMed
Drobizhev, M., Makarov, N.S., Tillo, S.E., Hughes, T.E. & Rebane, A. (2011). Two-photon absorption properties of fluorescent proteins. Nat Methods 8, 393–399.CrossRefGoogle ScholarPubMed
Horton, N.G., Wang, K., Kobat, D., Clark, C.G., Wise, F.W., Schaffer, C.B. & Xu, C. (2013). In vivo three-photon microscopy of subcortical structures within an intact mouse brain. Nature Photon 7, 205–209.CrossRefGoogle ScholarPubMed
Jayo, A., Parsons, M. & Adams, J.C. (2012). A novel rho-dependent pathway that drives interaction of fascin-1 with p-Lin-11/Isl-1/Mec-3 kinase (LIMK) 1/2 to promote fascin-1/actin binding and filopodia stability. BMC Biol 10, 72.CrossRefGoogle ScholarPubMed
Kobat, D., Durst, M.E., Nishimura, N., Wong, A.W., Schaffer, C.B. & Xu, C. (2009). Deep tissue multiphoton microscopy using longer wavelength excitation. Opt Express 17, 13354–13364.CrossRefGoogle ScholarPubMed
Lin, J. & Spence, D.J. (2016). 25.5 fs dissipative-soliton diamond Raman laser. Opt Lett 41, 1861–1864.CrossRefGoogle ScholarPubMed
Linnenbank, H. & Linden, S. (2014). High repetition rate femtosecond double pass optical parametric generator with more than 2 W tunable output in the NIR. Opt Express 22, 18072–18077.CrossRefGoogle ScholarPubMed
Mahou, P., Zimmerley, M., Loulier, K., Matho, K.S., Labroille, G., Morin, X., Supatto, W., Livet, J., Débarre, D. & Beaurepaire, E. (2012). Multicolor two-photon tissue imaging by wavelength mixing. Nat Methods 9, 815–818.CrossRefGoogle ScholarPubMed
Murtagh, M., Lin, J., Mildren, R.P., McConnell, G. & Spence, D.J. (2015a). Efficient diamond Raman laser generating 65 fs pulses. Opt Express 23, 15504–15513.CrossRefGoogle ScholarPubMed
Murtagh, M., Lin, J., Trägårdh, J., Mcconnell, G. & Spence, D.J. (2015b). Ultrafast second-Stokes diamond Raman laser. Opt Express 24, 8149–8155.CrossRefGoogle Scholar
Sun, C.K., Chu, S.W., Chen, S.Y., Tsai, T.H., Liu, T.M., Lin, C.Y. & Tsai, H.J. (2004). Higher harmonic generation microscopy for developmental biology. J Struct Biol 147, 19–30.CrossRefGoogle ScholarPubMed
Tillo, S.E., Hughes, T.E., Makarov, N.S., Rebane, A. & Drobizhev, M. (2010). A new approach to dual-color two-photon microscopy with fluorescent proteins. BMC Biotechnol. 10, 6.CrossRefGoogle ScholarPubMed
Trägårdh, J., Robb, G., Amor, R., Amos, W.B., Dempster, J. & McConnell, G. (2015). Exploration of the two-photon excitation spectrum of fluorescent dyes at wavelengths below the range of the Ti:Sapphire laser. J Microsc 259, 210–218.CrossRefGoogle ScholarPubMed
Xu, C. & Webb, W.W. (1996). Measurement of two-photon excitation cross sections of molecular fluorophores with data from 690 to 1050 nm. J Opt Soc Am B 13, 481–491.CrossRefGoogle Scholar
Xu, C., Zipfel, W., Shear, J.B., Williams, R.M. & Webb, W.W. (1996). Multiphoton fluorescence excitation: New spectral windows for biological nonlinear microscopy. Proc Natl Acad Sci USA 93, 10763–10768.CrossRefGoogle ScholarPubMed
Yamanaka, M., Saito, K., Smith, N.I., Arai, Y., Uegaki, K., Yonemaru, Y., Mochizuki, K., Kawata, S., Nagai, T. & Fujita, K. (2015). Visible-wavelength two-photon excitation microscopy for fluorescent protein imaging. J Biomed Opt 20, 101202.CrossRefGoogle ScholarPubMed
Zipfel, W.R., Williams, R.M. & Webb, W.W. (2003). Nonlinear magic: Multiphoton microscopy in the biosciences. Nat Biotechnol 21, 1369–1377.CrossRefGoogle ScholarPubMed