Abstract

The video stream captured by an in-line holographic microscope can be analyzed on a frame-by-frame basis to track individual colloidal particles’ three-dimensional motions with nanometer resolution, and simultaneously to measure their sizes and refractive indexes. Through a combination of hardware acceleration and software optimization, this analysis can be carried out in near real time with off-the-shelf instrumentation. An efficient particle identification algorithm automates initial position estimation with sufficient accuracy to enable unattended holographic tracking and characterization. This technique’s resolution for particle size is fine enough to detect molecular-scale coatings on the surfaces of colloidal spheres, without requiring staining or fluorescent labeling. We demonstrate this approach to label-free holographic flow cytometry by detecting the binding of avidin to biotinylated polystyrene spheres.

© 2009 Optical Society of America

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References

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  1. S.-H. Lee, Y. Roichman, G.-R. Yi, S.-H. Kim, S.-M. Yang, A. van Blaaderen, P. van Oostrum, D. G. Grier, “Characterizing and tracking single colloidal particles with video holographic microscopy,” Opt. Express 15, 18275–18282 (2007), http://www.opticsinfobase.org/oe/abstract.cfm?URI=oe-15-26-18275.
    [CrossRef] [PubMed]
  2. J. Sheng, E. Malkiel, J. Katz, “Digital holographic microscope for measuring three-dimensional particle distributions and motions,” Appl. Opt. 45(16), 3893–3901 (2006).
    [CrossRef]
  3. S.-H. Lee, D. G. Grier, “Holographic microscopy of holographically trapped three-dimensional structures,” Opt. Express 15, 1505–1512 (2007), http://www.opticsinfobase.org/oe/abstract.cfm?URI=oe-15-4-1505.
    [CrossRef] [PubMed]
  4. T. Savin, P. S. Doyle, “Role of finite exposure time on measuring an elastic modulus using microrheology,” Phys. Rev. E 71, 041106 (2005).
    [CrossRef]
  5. T. Savin, P. S. Doyle, “Static and dynamic errors in particle tracking microrheology,” Biophys. J. 88, 623–638 (2005).
    [CrossRef]
  6. C. F. Bohren, D. R. Huffman, Absorption and Scattering of Light by Small Particles (Wiley Interscience, New York, 1983).
  7. J. C. Crocker, D. G. Grier, “Methods of digital video microscopy for colloidal studies,” J. Colloid Interface Sci. 179, 298–310 (1996).
    [CrossRef]
  8. Y. Roichman, B. Sun, A. Stolarski, D. G. Grier, “Influence of non-conservative optical forces on the dynamics of optically trapped colloidal spheres: The fountain of probability,” Phys. Rev. Lett. 101, 128301 (2008).
    [PubMed]
  9. F. C. Cheong, S. Duarte, S.-H. Lee, D. G. Grier, “Holographic microrheology of polysaccharides from Streptococcus mutans biofilms,” Rheologica Acta 48, 109–115 (2009).
    [CrossRef]
  10. F. C. Cheong, K. Xiao, D. G. Grier, “Characterization of individual milk fat globules with holographic video microscopy,” J. Dairy Sci. 92, 95–99 (2009).
    [CrossRef]
  11. J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
    [CrossRef]
  12. C. B. Markwardt, “Non-linear least squares fitting in IDL with MPFIT,” in Astronomical Data Analysis Software and Systems XVIII, D. Bohlender, P. Dowler, D. Durand, eds. (Astronomical Society of the Pacific, San Francisco, 2009), in press.
  13. P. Messmer, P. J. Mullowney, B. E. Granger, “GPULib: GPU computing in high-level languages,” Comp. Sci. Eng. 10, 70–73 (2008).
    [CrossRef]
  14. R. O. Duda, P. E. Hart, “Use of the Hough transformation to detect lines and curves in pictures,” Commun. ACM 15, 11–15 (1972).
    [CrossRef]
  15. X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
    [CrossRef]

2009

F. C. Cheong, S. Duarte, S.-H. Lee, D. G. Grier, “Holographic microrheology of polysaccharides from Streptococcus mutans biofilms,” Rheologica Acta 48, 109–115 (2009).
[CrossRef]

F. C. Cheong, K. Xiao, D. G. Grier, “Characterization of individual milk fat globules with holographic video microscopy,” J. Dairy Sci. 92, 95–99 (2009).
[CrossRef]

2008

P. Messmer, P. J. Mullowney, B. E. Granger, “GPULib: GPU computing in high-level languages,” Comp. Sci. Eng. 10, 70–73 (2008).
[CrossRef]

2007

2006

J. Sheng, E. Malkiel, J. Katz, “Digital holographic microscope for measuring three-dimensional particle distributions and motions,” Appl. Opt. 45(16), 3893–3901 (2006).
[CrossRef]

2005

T. Savin, P. S. Doyle, “Role of finite exposure time on measuring an elastic modulus using microrheology,” Phys. Rev. E 71, 041106 (2005).
[CrossRef]

T. Savin, P. S. Doyle, “Static and dynamic errors in particle tracking microrheology,” Biophys. J. 88, 623–638 (2005).
[CrossRef]

2003

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

1996

J. C. Crocker, D. G. Grier, “Methods of digital video microscopy for colloidal studies,” J. Colloid Interface Sci. 179, 298–310 (1996).
[CrossRef]

1972

R. O. Duda, P. E. Hart, “Use of the Hough transformation to detect lines and curves in pictures,” Commun. ACM 15, 11–15 (1972).
[CrossRef]

Bohren, C. F.

C. F. Bohren, D. R. Huffman, Absorption and Scattering of Light by Small Particles (Wiley Interscience, New York, 1983).

Brock, R. S.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Cheong, F. C.

F. C. Cheong, K. Xiao, D. G. Grier, “Characterization of individual milk fat globules with holographic video microscopy,” J. Dairy Sci. 92, 95–99 (2009).
[CrossRef]

F. C. Cheong, S. Duarte, S.-H. Lee, D. G. Grier, “Holographic microrheology of polysaccharides from Streptococcus mutans biofilms,” Rheologica Acta 48, 109–115 (2009).
[CrossRef]

Crocker, J. C.

J. C. Crocker, D. G. Grier, “Methods of digital video microscopy for colloidal studies,” J. Colloid Interface Sci. 179, 298–310 (1996).
[CrossRef]

Doyle, P. S.

T. Savin, P. S. Doyle, “Static and dynamic errors in particle tracking microrheology,” Biophys. J. 88, 623–638 (2005).
[CrossRef]

T. Savin, P. S. Doyle, “Role of finite exposure time on measuring an elastic modulus using microrheology,” Phys. Rev. E 71, 041106 (2005).
[CrossRef]

Duarte, S.

F. C. Cheong, S. Duarte, S.-H. Lee, D. G. Grier, “Holographic microrheology of polysaccharides from Streptococcus mutans biofilms,” Rheologica Acta 48, 109–115 (2009).
[CrossRef]

Duda, R. O.

R. O. Duda, P. E. Hart, “Use of the Hough transformation to detect lines and curves in pictures,” Commun. ACM 15, 11–15 (1972).
[CrossRef]

Govinadaraju, N.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Granger, B. E.

P. Messmer, P. J. Mullowney, B. E. Granger, “GPULib: GPU computing in high-level languages,” Comp. Sci. Eng. 10, 70–73 (2008).
[CrossRef]

Grier, D. G.

F. C. Cheong, S. Duarte, S.-H. Lee, D. G. Grier, “Holographic microrheology of polysaccharides from Streptococcus mutans biofilms,” Rheologica Acta 48, 109–115 (2009).
[CrossRef]

F. C. Cheong, K. Xiao, D. G. Grier, “Characterization of individual milk fat globules with holographic video microscopy,” J. Dairy Sci. 92, 95–99 (2009).
[CrossRef]

S.-H. Lee, Y. Roichman, G.-R. Yi, S.-H. Kim, S.-M. Yang, A. van Blaaderen, P. van Oostrum, D. G. Grier, “Characterizing and tracking single colloidal particles with video holographic microscopy,” Opt. Express 15, 18275–18282 (2007), http://www.opticsinfobase.org/oe/abstract.cfm?URI=oe-15-26-18275.
[CrossRef] [PubMed]

S.-H. Lee, D. G. Grier, “Holographic microscopy of holographically trapped three-dimensional structures,” Opt. Express 15, 1505–1512 (2007), http://www.opticsinfobase.org/oe/abstract.cfm?URI=oe-15-4-1505.
[CrossRef] [PubMed]

J. C. Crocker, D. G. Grier, “Methods of digital video microscopy for colloidal studies,” J. Colloid Interface Sci. 179, 298–310 (1996).
[CrossRef]

Y. Roichman, B. Sun, A. Stolarski, D. G. Grier, “Influence of non-conservative optical forces on the dynamics of optically trapped colloidal spheres: The fountain of probability,” Phys. Rev. Lett. 101, 128301 (2008).
[PubMed]

Harris, M.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Hart, P. E.

R. O. Duda, P. E. Hart, “Use of the Hough transformation to detect lines and curves in pictures,” Commun. ACM 15, 11–15 (1972).
[CrossRef]

Hu, X.-H.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Huffman, D. R.

C. F. Bohren, D. R. Huffman, Absorption and Scattering of Light by Small Particles (Wiley Interscience, New York, 1983).

Jacobs, K. M.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Katz, J.

J. Sheng, E. Malkiel, J. Katz, “Digital holographic microscope for measuring three-dimensional particle distributions and motions,” Appl. Opt. 45(16), 3893–3901 (2006).
[CrossRef]

Kim, S.-H.

Kruger, J.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Lee, S.-H.

Lefohn, A. E.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Lu, J. Q.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Luebke, D.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Ma, X.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Malkiel, E.

J. Sheng, E. Malkiel, J. Katz, “Digital holographic microscope for measuring three-dimensional particle distributions and motions,” Appl. Opt. 45(16), 3893–3901 (2006).
[CrossRef]

Markwardt, C. B.

C. B. Markwardt, “Non-linear least squares fitting in IDL with MPFIT,” in Astronomical Data Analysis Software and Systems XVIII, D. Bohlender, P. Dowler, D. Durand, eds. (Astronomical Society of the Pacific, San Francisco, 2009), in press.

Messmer, P.

P. Messmer, P. J. Mullowney, B. E. Granger, “GPULib: GPU computing in high-level languages,” Comp. Sci. Eng. 10, 70–73 (2008).
[CrossRef]

Mullowney, P. J.

P. Messmer, P. J. Mullowney, B. E. Granger, “GPULib: GPU computing in high-level languages,” Comp. Sci. Eng. 10, 70–73 (2008).
[CrossRef]

Owens, J. D.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Purcell, T. J.

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Roichman, Y.

Savin, T.

T. Savin, P. S. Doyle, “Role of finite exposure time on measuring an elastic modulus using microrheology,” Phys. Rev. E 71, 041106 (2005).
[CrossRef]

T. Savin, P. S. Doyle, “Static and dynamic errors in particle tracking microrheology,” Biophys. J. 88, 623–638 (2005).
[CrossRef]

Sheng, J.

J. Sheng, E. Malkiel, J. Katz, “Digital holographic microscope for measuring three-dimensional particle distributions and motions,” Appl. Opt. 45(16), 3893–3901 (2006).
[CrossRef]

Stolarski, A.

Y. Roichman, B. Sun, A. Stolarski, D. G. Grier, “Influence of non-conservative optical forces on the dynamics of optically trapped colloidal spheres: The fountain of probability,” Phys. Rev. Lett. 101, 128301 (2008).
[PubMed]

Sun, B.

Y. Roichman, B. Sun, A. Stolarski, D. G. Grier, “Influence of non-conservative optical forces on the dynamics of optically trapped colloidal spheres: The fountain of probability,” Phys. Rev. Lett. 101, 128301 (2008).
[PubMed]

van Blaaderen, A.

van Oostrum, P.

Xiao, K.

F. C. Cheong, K. Xiao, D. G. Grier, “Characterization of individual milk fat globules with holographic video microscopy,” J. Dairy Sci. 92, 95–99 (2009).
[CrossRef]

Yang, P.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Yang, S.-M.

Yi, G.-R.

Appl. Opt.

J. Sheng, E. Malkiel, J. Katz, “Digital holographic microscope for measuring three-dimensional particle distributions and motions,” Appl. Opt. 45(16), 3893–3901 (2006).
[CrossRef]

Biophys. J.

T. Savin, P. S. Doyle, “Static and dynamic errors in particle tracking microrheology,” Biophys. J. 88, 623–638 (2005).
[CrossRef]

Commun. ACM

R. O. Duda, P. E. Hart, “Use of the Hough transformation to detect lines and curves in pictures,” Commun. ACM 15, 11–15 (1972).
[CrossRef]

Comp. Graph. Forum

J. D. Owens, D. Luebke, N. Govinadaraju, M. Harris, J. Kruger, A. E. Lefohn, T. J. Purcell, “A survey of general-purpose computation on graphics hardware,” Comp. Graph. Forum 26, 80–113 (2007).
[CrossRef]

Comp. Sci. Eng.

P. Messmer, P. J. Mullowney, B. E. Granger, “GPULib: GPU computing in high-level languages,” Comp. Sci. Eng. 10, 70–73 (2008).
[CrossRef]

J. Colloid Interface Sci.

J. C. Crocker, D. G. Grier, “Methods of digital video microscopy for colloidal studies,” J. Colloid Interface Sci. 179, 298–310 (1996).
[CrossRef]

J. Dairy Sci.

F. C. Cheong, K. Xiao, D. G. Grier, “Characterization of individual milk fat globules with holographic video microscopy,” J. Dairy Sci. 92, 95–99 (2009).
[CrossRef]

Opt. Express

Phys. Med. Bio.

X. Ma, J. Q. Lu, R. S. Brock, K. M. Jacobs, P. Yang, X.-H. Hu, “Determination of complex refractive index of polystyrene microspheres from 370 to 1610 nm,” Phys. Med. Bio. 48, 4165–4172 (2003).
[CrossRef]

Phys. Rev. E

T. Savin, P. S. Doyle, “Role of finite exposure time on measuring an elastic modulus using microrheology,” Phys. Rev. E 71, 041106 (2005).
[CrossRef]

Phys. Rev. Lett.

Y. Roichman, B. Sun, A. Stolarski, D. G. Grier, “Influence of non-conservative optical forces on the dynamics of optically trapped colloidal spheres: The fountain of probability,” Phys. Rev. Lett. 101, 128301 (2008).
[PubMed]

Rheologica Acta

F. C. Cheong, S. Duarte, S.-H. Lee, D. G. Grier, “Holographic microrheology of polysaccharides from Streptococcus mutans biofilms,” Rheologica Acta 48, 109–115 (2009).
[CrossRef]

Other

C. F. Bohren, D. R. Huffman, Absorption and Scattering of Light by Small Particles (Wiley Interscience, New York, 1983).

C. B. Markwardt, “Non-linear least squares fitting in IDL with MPFIT,” in Astronomical Data Analysis Software and Systems XVIII, D. Bohlender, P. Dowler, D. Durand, eds. (Astronomical Society of the Pacific, San Francisco, 2009), in press.

Supplementary Material (1)

» Media 1: MOV (442 KB)     

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Figures (5)

Fig. 1.
Fig. 1.

In-line holographic video microscope. A collimated laser beam illuminates the sample. Light scattered by the sample interferes with the unscattered portion of the beam in the focal plane of the objective lens. The interference pattern is magnified, recorded and then fit to predictions of Lorenz-Mie theory to obtain measurements of the particle’s position, its size, and its refractive index.

Fig. 2.
Fig. 2.

Detecting particle images in a video hologram. Original (a) and transformed (b) holographic images of three colloidal spheres. Superimposed line segments in (a) indicate the votes cast by three representative pixels. Intensity in (b) is scaled by the number of votes, with black representing 0 and white representing 800 votes. Superimposed surface plots illustrate the middle sphere’s transformation. Scale bar indicates 10 µm. Media 1 shows detection, identification and fitting of a moving particle in a holographic video sequence.

Fig. 3.
Fig. 3.

Holographic particle-image velocimetry. (a) Measured three-dimensional trajectories of 500 colloidal spheres travelling down a microfluidic channel in a pressure-driven flow. Each sphere represents the position of a particle in one field of a holographic snapshot. Features from a sequence of fields are linked into trajectories that are colored by the particle’s measured speed. (b) Poiseuille flow profile along the vertical direction obtained from the data in (a). Particles are excluded from the shaded regions by their interactions with the upper and lower glass walls of the channel. The dashed curve is a fit to the anticipated parabolic flow profile.

Fig. 4.
Fig. 4.

Holographic characterization of streaming particles. (a) Trajectory-averaged radii ap and refractive indexes np for a sample of commercial polystyrene spheres in water. Histograms show the distributions of observed sizes and refractive indexes, together with Gaussian fits. (b) Trajectory-averaged radius and refractive index as a function of mean speed.

Fig. 5.
Fig. 5.

Detection of avidin binding to biotinylated polystyrene spheres. (a) Yellow circles show the probability distribution for the measured particle radii in stock biotinylated polystyrene spheres. Red circles show the corresponding distribution for a sample of these spheres after incubation with neutravidin. Dashed curves are guides to the eye. (b) Equivalent distributions for particles’ refractive indexes. Arrow indicates redistribution of probability from low density tail in the stock sample to the peak in the coated sample.

Equations (1)

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I ( r ) = E 0 ( z z p ) + E s ( r r p ) 2 ,

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