Abstract

We present a two-photon microscope that is approximately 2.9g in mass and 2.0×1.9×1.1cm3 in size and based on a microelectromechanical systems (MEMS) laser-scanning mirror. The microscope has a focusing motor and a micro-optical assembly composed of four gradient refractive index lenses and a dichroic microprism. Fluorescence is captured without the detected emissions reflecting off the MEMS mirror, by use of separate optical fibers for fluorescence collection and delivery of ultrashort excitation pulses. Using this microscope we imaged neocortical microvasculature and tracked the flow of erythrocytes in live mice.

© 2009 Optical Society of America

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2008 (9)

2007 (1)

J. Sawinski and W. Denk, J. Appl. Phys. 102, 034701 (2007).
[CrossRef]

2006 (4)

2005 (1)

2004 (2)

2001 (1)

F. Helmchen, M. S. Fee, D. W. Tank, and W. Denk, Neuron 31, 903 (2001).
[CrossRef] [PubMed]

1998 (1)

D. Kleinfeld, P. P. Mitra, F. Helmchen, and W. Denk, Proc. Natl. Acad. Sci. USA 95, 15741 (1998).
[CrossRef] [PubMed]

Ahn, Y. C.

Allen, J.

Anderson, E. P.

Bao, H.

Barretto, R. P.

M. E. Llewellyn, R. P. Barretto, S. L. Delp, and M. J. Schnitzer, Nature 454, 784 (2008).
[PubMed]

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

W. Piyawattanametha, R. P. Barretto, T. H. Ko, B. A. Flusberg, E. D. Cocker, H. Ra, D. Lee, O. Solgaard, and M. J. Schnitzer, Opt. Lett. 31, 2018 (2006).
[CrossRef] [PubMed]

Ben-Yakar, A.

Burns, L. D.

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

Chan, M. C.

Chen, I. H.

Chen, P.

Chen, Z.

Chia, S. H.

Cocker, E. D.

Cranfield, C.

Delp, S. L.

M. E. Llewellyn, R. P. Barretto, S. L. Delp, and M. J. Schnitzer, Nature 454, 784 (2008).
[PubMed]

Denk, W.

J. Sawinski and W. Denk, J. Appl. Phys. 102, 034701 (2007).
[CrossRef]

F. Helmchen, M. S. Fee, D. W. Tank, and W. Denk, Neuron 31, 903 (2001).
[CrossRef] [PubMed]

D. Kleinfeld, P. P. Mitra, F. Helmchen, and W. Denk, Proc. Natl. Acad. Sci. USA 95, 15741 (1998).
[CrossRef] [PubMed]

Durr, N. J.

Engelbrecht, C. J.

Evans, C.

Fee, M. S.

F. Helmchen, M. S. Fee, D. W. Tank, and W. Denk, Neuron 31, 903 (2001).
[CrossRef] [PubMed]

Flusberg, B. A.

Freudiger, C. W.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Fu, L.

Ganikhanov, F.

Göbel, W.

Gu, M.

He, C.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Helmchen, F.

Holtom, G. R.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Hoy, C. L.

Jain, A.

Johnston, R. S.

Jung, J. C.

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

B. A. Flusberg, J. C. Jung, E. D. Cocker, E. P. Anderson, and M. J. Schnitzer, Opt. Lett. 30, 2272 (2005).
[CrossRef] [PubMed]

Jung, W.

Kang, J. X.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Kerr, J. N.

Kleinfeld, D.

D. Kleinfeld, P. P. Mitra, F. Helmchen, and W. Denk, Proc. Natl. Acad. Sci. USA 95, 15741 (1998).
[CrossRef] [PubMed]

Ko, T. H.

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

W. Piyawattanametha, R. P. Barretto, T. H. Ko, B. A. Flusberg, E. D. Cocker, H. Ra, D. Lee, O. Solgaard, and M. J. Schnitzer, Opt. Lett. 31, 2018 (2006).
[CrossRef] [PubMed]

König, K.

Krasieva, T. B.

Le Harzic, R.

Lee, D.

Legare, F.

Li, X.

Liu, T. M.

Llewellyn, M. E.

M. E. Llewellyn, R. P. Barretto, S. L. Delp, and M. J. Schnitzer, Nature 454, 784 (2008).
[PubMed]

Lu, S.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

MacDonald, D. J.

McCormic, D. T.

Messerschmidt, B.

Min, W.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Mitra, P. P.

D. Kleinfeld, P. P. Mitra, F. Helmchen, and W. Denk, Proc. Natl. Acad. Sci. USA 95, 15741 (1998).
[CrossRef] [PubMed]

Mukamel, E. A.

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

Myaing, M. T.

Nimmerjahn, A.

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

W. Göbel, A. Nimmerjahn, and F. Helmchen, Opt. Lett. 29, 1285 (2004).
[CrossRef] [PubMed]

W. Göbel, J. N. Kerr, A. Nimmerjahn, and F. Helmchen, Opt. Lett. 29, 2521 (2004).
[CrossRef] [PubMed]

Pattie, R.

Piyawattanametha, W.

Ra, H.

Riemann, I.

Saar, B. G.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Sawinski, J.

J. Sawinski and W. Denk, J. Appl. Phys. 102, 034701 (2007).
[CrossRef]

Schnitzer, M. J.

M. E. Llewellyn, R. P. Barretto, S. L. Delp, and M. J. Schnitzer, Nature 454, 784 (2008).
[PubMed]

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

W. Piyawattanametha, R. P. Barretto, T. H. Ko, B. A. Flusberg, E. D. Cocker, H. Ra, D. Lee, O. Solgaard, and M. J. Schnitzer, Opt. Lett. 31, 2018 (2006).
[CrossRef] [PubMed]

B. A. Flusberg, J. C. Jung, E. D. Cocker, E. P. Anderson, and M. J. Schnitzer, Opt. Lett. 30, 2272 (2005).
[CrossRef] [PubMed]

Seibel, E. J.

Solgaard, O.

Su, J.

Sun, C. K.

Tang, S.

Tank, D. W.

F. Helmchen, M. S. Fee, D. W. Tank, and W. Denk, Neuron 31, 903 (2001).
[CrossRef] [PubMed]

Tomov, I. V.

Tromberg, B. J.

Tsai, J. C.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

Vance, R.

Weinigel, M.

Xie, H.

Xie, T.

Xie, X. S.

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

F. Legare, C. Evans, F. Ganikhanov, and X. S. Xie, Opt. Express 14, 4427 (2006).
[CrossRef] [PubMed]

J. Appl. Phys. (1)

J. Sawinski and W. Denk, J. Appl. Phys. 102, 034701 (2007).
[CrossRef]

Nat. Methods (1)

B. A. Flusberg, A. Nimmerjahn, E. D. Cocker, E. A. Mukamel, R. P. Barretto, T. H. Ko, L. D. Burns, J. C. Jung, and M. J. Schnitzer, Nat. Methods 5, 935 (2008).
[CrossRef] [PubMed]

Nature (1)

M. E. Llewellyn, R. P. Barretto, S. L. Delp, and M. J. Schnitzer, Nature 454, 784 (2008).
[PubMed]

Neuron (1)

F. Helmchen, M. S. Fee, D. W. Tank, and W. Denk, Neuron 31, 903 (2001).
[CrossRef] [PubMed]

Opt. Express (6)

Opt. Lett. (7)

Proc. Natl. Acad. Sci. USA (1)

D. Kleinfeld, P. P. Mitra, F. Helmchen, and W. Denk, Proc. Natl. Acad. Sci. USA 95, 15741 (1998).
[CrossRef] [PubMed]

Science (1)

C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. He, J. C. Tsai, J. X. Kang, and X. S. Xie, Science 322, 1857 (2008).
[CrossRef] [PubMed]

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Figures (3)

Fig. 1
Fig. 1

a, Portable two-photon microscope. Electrical lines control the MEMS scanner and focusing micromotor. b, The scanner die is wirebonded onto electrodes on the printed circuit board (PCB). c, Computer-aided-design model of the microscope, in a cut-away view. d, Laser illumination (red arrows) and fluorescence collection (green arrows) pathways.

Fig. 2
Fig. 2

a, Optical deflection angle versus dc voltage, and b, frequency response function for inner (blue solid curve) and outer (red dashed curve) axes of a 1 mm × 1 mm MEMS scanner. For each axis time-varying signals were applied to one of the two opposing comb banks. c, Lateral and d, axial resolutions were determined as the FWHM of curve fits (Airy function squared, c, and Gaussian, d) to normalized, cross-sectional images of 100 nm diameter fluorescent beads. Data from three beads (circle, square, and diamond symbols) and fits from one bead (solid curves) are shown.

Fig. 3
Fig. 3

a–c, Images of neocortical capillaries, averaged over eight frames acquired over 2 s at 4 Hz . d–f, Line images taken by driving only the scanner’s outer axis, at its resonant frequency ( 560 Hz ) . Flowing erythrocytes appear as dark streaks in relief. Flow velocities were found from the slopes of the dark streaks at the central region of the line scan. Illumination power was 27 mW at the sample. Scale bar in a also applies to b and c. Scale bar in e also applies to f.

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