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Optics Express

Optics Express

  • Editor: Andrew M. Weiner
  • Vol. 21, Iss. 14 — Jul. 15, 2013
  • pp: 17256–17264

Two-photon imaging of multiple fluorescent proteins by phase-shaping and linear unmixing with a single broadband laser

Meredith H. Brenner, Dawen Cai, Joel A. Swanson, and Jennifer P. Ogilvie  »View Author Affiliations


Optics Express, Vol. 21, Issue 14, pp. 17256-17264 (2013)
http://dx.doi.org/10.1364/OE.21.017256


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Abstract

Imaging multiple fluorescent proteins (FPs) by two-photon microscopy has numerous applications for studying biological processes in thick and live samples. Here we demonstrate a setup utilizing a single broadband laser and a phase-only pulse-shaper to achieve imaging of three FPs (mAmetrine, TagRFPt, and mKate2) in live mammalian cells. Phase-shaping to achieve selective excitation of the FPs in combination with post-imaging linear unmixing enables clean separation of the fluorescence signal of each FP. This setup also benefits from low overall cost and simple optical alignment, enabling easy adaptation in a regular biomedical research laboratory.

© 2013 OSA

OCIS Codes
(180.2520) Microscopy : Fluorescence microscopy
(190.4180) Nonlinear optics : Multiphoton processes
(320.5540) Ultrafast optics : Pulse shaping
(180.4315) Microscopy : Nonlinear microscopy

ToC Category:
Microscopy

History
Original Manuscript: May 28, 2013
Revised Manuscript: July 3, 2013
Manuscript Accepted: July 5, 2013
Published: July 11, 2013

Virtual Issues
Vol. 8, Iss. 8 Virtual Journal for Biomedical Optics

Citation
Meredith H. Brenner, Dawen Cai, Joel A. Swanson, and Jennifer P. Ogilvie, "Two-photon imaging of multiple fluorescent proteins by phase-shaping and linear unmixing with a single broadband laser," Opt. Express 21, 17256-17264 (2013)
http://www.opticsinfobase.org/oe/abstract.cfm?URI=oe-21-14-17256


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