Abstract

Autofocus is a critical aspect of any automated microscopy application. Here we report on the successful exploitation of chromatic aberration to speed autofocus for biological microscopy. Using pairs of lenses aligned nominally as infinite conjugates, the chromatic aberration produced by a microscope can be manipulated to achieve multiplanar imaging covering a wide range of spacings. Using both three-chip CCD and Bayer-array color cameras, chromatic aberration-based multiplanar imaging is used to reduce by a factor of 3 the number of mechanical movements of the biological sample necessary to determine best focus. Chromatic aberration-based autofocus is validated using a 20× 0.75NA objective while scanning over 6000 fields of National Institutes of Health 3T3 cells cultured on a coverglass. The combined precisions (standard deviations) of 23.7nm for a three-chip CCD camera and 23.3nm for a Bayer- array camera are obtained. These precisions are approximately 1/20th of the depth of field of the objective.

© 2011 Optical Society of America

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2009 (2)

M. Weinigel, A. L. Kellner, and J. H. Price, “Exploration of chromatic aberration for multiplanar imaging: proof of concept with implications for fast, efficient autofocus,” Cytometry A 75A, 999–1006 (2009).
[CrossRef]

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

2008 (1)

2007 (2)

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

2006 (1)

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

2005 (1)

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

2004 (1)

2000 (3)

S. Cha, P. C. Lin, L. Zhu, P.-C. Sun, and Y. Fainman, “Nontranslational three-dimensional profilometry by chromatic confocal microscopy with dynamically configurable micromirror scanning,” Appl. Opt. 39, 2605–2613 (2000).
[CrossRef]

L. K. Nguyen, M. E. Bravo-Zanoguera, A. L. Kellner, and J. H. Price, “Magnification corrected optical image splitting module for simultaneous multiplanar acquisition,” Proc. SPIE 3921, 31–40 (2000).
[CrossRef]

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

1999 (1)

1998 (2)

P. C. Lin, P.-C. Sun, L. Zhu, and Y. Fainman, “Single-shot depth-section imaging through chromatic slit-scan confocal microscopy,” Appl. Opt. 37, 6764–6770 (1998).
[CrossRef]

M. Bravo-Zanoguera, B. von Massenbach, A. L. Kellner, and J. H. Price, “High-performance autofocus circuit for biological microscopy,” Rev. Sci. Instrum. 69, 3966–3977 (1998).
[CrossRef]

1997 (1)

1994 (3)

M. Maly and A. Boyde, “Real-time stereoscopic confocal reflection microscopy using objective lenses with linear longitudinal chromatic dispersion,” Scanning 16, 187–192 (1994).

H. J. Tiziani and H.-M. Uhde, “Three-dimensional image sensing by chromatic confocal microscopy,” Appl. Opt. 33, 1838–1843 (1994).
[CrossRef] [PubMed]

J. H. Price and D. A. Gough, “Comparison of phase-contrast and fluorescence digital autofocus for scanning microscopy,” Cytometry 16, 283–297 (1994).
[CrossRef] [PubMed]

1992 (1)

M. A. Browne, O. Akinyemi, and A. Boyde, “Confocal surface profiling utilizing chromatic aberration,” Scanning 14, 145–153 (1992).
[CrossRef]

1985 (1)

F. C. A. Groen, I. T. Young, and G. Ligthart, “A comparison of different focus functions for use in autofocus algorithms,” Cytometry 6, 81–91 (1985).
[CrossRef] [PubMed]

1984 (1)

G. Molesini, G. Pedrini, P. Poggi, and F. Quercioli, “Focus-wavelength encoded optical profilometer,” Opt. Commun. 49, 229–233 (1984).
[CrossRef]

Agustin, R. M.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

Akinyemi, O.

M. A. Browne, O. Akinyemi, and A. Boyde, “Confocal surface profiling utilizing chromatic aberration,” Scanning 14, 145–153 (1992).
[CrossRef]

Akong, M.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Bach, H.

H. Bach and N. Neuroth, The Properties of Optical Glass (Springer-Verlag, 1998).

Born, M.

M. Born and E. Wolf, Principles of Optics (Pergamon, 1980), 6th ed.

Boyde, A.

M. Maly and A. Boyde, “Real-time stereoscopic confocal reflection microscopy using objective lenses with linear longitudinal chromatic dispersion,” Scanning 16, 187–192 (1994).

M. A. Browne, O. Akinyemi, and A. Boyde, “Confocal surface profiling utilizing chromatic aberration,” Scanning 14, 145–153 (1992).
[CrossRef]

Bravo-Zanoguera, M.

M. A. Oliva, M. Bravo-Zanoguera, and J. H. Price, “Filtering out contrast reversals for microscopy autofocus,” Appl. Opt. 38, 638–646 (1999).
[CrossRef]

M. Bravo-Zanoguera, B. von Massenbach, A. L. Kellner, and J. H. Price, “High-performance autofocus circuit for biological microscopy,” Rev. Sci. Instrum. 69, 3966–3977 (1998).
[CrossRef]

Bravo-Zanoguera, M. E.

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

L. K. Nguyen, M. E. Bravo-Zanoguera, A. L. Kellner, and J. H. Price, “Magnification corrected optical image splitting module for simultaneous multiplanar acquisition,” Proc. SPIE 3921, 31–40 (2000).
[CrossRef]

Browne, M. A.

M. A. Browne, O. Akinyemi, and A. Boyde, “Confocal surface profiling utilizing chromatic aberration,” Scanning 14, 145–153 (1992).
[CrossRef]

Buehrer, B. M.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

Callaway, S.

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Cha, S.

Chemerisov, S.

Cheng, P.-C.

P.-C. Cheng, “The contrast formation in optical microscopy,” in Handbook of Biological Confocal Microscopy, 3rd ed. (Springer, 2006), pp. 162–206.
[CrossRef]

DeLeon, R. P.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Dobson, S. L.

Fainman, Y.

Fox, M. D. T.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Françon, M.

M. Françon, Progress in Microscopy, Vol.  9 of International Series of Monographs on Pure and Applied Biology(Pergamon, 1961).

Geiger, B.

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

Goodacre, A.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Gough, D. A.

J. H. Price and D. A. Gough, “Comparison of phase-contrast and fluorescence digital autofocus for scanning microscopy,” Cytometry 16, 283–297 (1994).
[CrossRef] [PubMed]

Groen, F. C. A.

F. C. A. Groen, I. T. Young, and G. Ligthart, “A comparison of different focus functions for use in autofocus algorithms,” Cytometry 6, 81–91 (1985).
[CrossRef] [PubMed]

Hand, D. P.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Haran, F. M.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Heynen, S.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Hunter, E. A.

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Ingermanson, R. S.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

Inoué, S.

S. Inoué and K. R. Spring, Video Microscopy: The Fundamentals, 2nd ed. (Springer, 1997).
[CrossRef]

Johnson, M. K.

M. K. Johnson, “Lighting and optical tools for image forensics,” Ph.D. thesis (Dartmouth College, 2007).

Jones, J. D. C.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Kam, Z.

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

Kellner, A. L.

M. Weinigel, A. L. Kellner, and J. H. Price, “Exploration of chromatic aberration for multiplanar imaging: proof of concept with implications for fast, efficient autofocus,” Cytometry A 75A, 999–1006 (2009).
[CrossRef]

L. K. Nguyen, M. E. Bravo-Zanoguera, A. L. Kellner, and J. H. Price, “Magnification corrected optical image splitting module for simultaneous multiplanar acquisition,” Proc. SPIE 3921, 31–40 (2000).
[CrossRef]

M. Bravo-Zanoguera, B. von Massenbach, A. L. Kellner, and J. H. Price, “High-performance autofocus circuit for biological microscopy,” Rev. Sci. Instrum. 69, 3966–3977 (1998).
[CrossRef]

Laris, C.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Laris, C. A.

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

Li, P.

Li, Y.

Ligthart, G.

F. C. A. Groen, I. T. Young, and G. Ligthart, “A comparison of different focus functions for use in autofocus algorithms,” Cytometry 6, 81–91 (1985).
[CrossRef] [PubMed]

Lin, P. C.

Liron, Y.

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

Liu, Z.

Loy, P. A.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

Maly, M.

M. Maly and A. Boyde, “Real-time stereoscopic confocal reflection microscopy using objective lenses with linear longitudinal chromatic dispersion,” Scanning 16, 187–192 (1994).

Mancini, M. A.

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

McDonough, P. M.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

McLean, M. A.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Mikic, I.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Molesini, G.

G. Molesini, G. Pedrini, P. Poggi, and F. Quercioli, “Focus-wavelength encoded optical profilometer,” Opt. Commun. 49, 229–233 (1984).
[CrossRef]

Moran, T. J.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Morelock, M. M.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Morgan, S. A.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Neuroth, N.

H. Bach and N. Neuroth, The Properties of Optical Glass (Springer-Verlag, 1998).

Nguyen, L. K.

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

L. K. Nguyen, M. E. Bravo-Zanoguera, A. L. Kellner, and J. H. Price, “Magnification corrected optical image splitting module for simultaneous multiplanar acquisition,” Proc. SPIE 3921, 31–40 (2000).
[CrossRef]

L. K. Nguyen, “High-throughput image cytometer for detection of circulating tumor cells and contrast-enhancement filtering for automated 3D image segmentation of cartilage tissue explants,” Ph.D. thesis (University of California–San Diego, 2007).

Nicoll, J. B.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

Oliva, M.

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

Oliva, M. A.

Paran, Y.

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

Pedrini, G.

G. Molesini, G. Pedrini, P. Poggi, and F. Quercioli, “Focus-wavelength encoded optical profilometer,” Opt. Commun. 49, 229–233 (1984).
[CrossRef]

Peters, C.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Peters, J.

J. Peters, “Nikon Instruments TiE-PFS dynamic focusing system,” Nature Methods—Application Notes, http://www.nature.com/app_notes/nmeth/2008/082312/full/an6676.html (2008).

Poggi, P.

G. Molesini, G. Pedrini, P. Poggi, and F. Quercioli, “Focus-wavelength encoded optical profilometer,” Opt. Commun. 49, 229–233 (1984).
[CrossRef]

Price, J. H.

M. Weinigel, A. L. Kellner, and J. H. Price, “Exploration of chromatic aberration for multiplanar imaging: proof of concept with implications for fast, efficient autofocus,” Cytometry A 75A, 999–1006 (2009).
[CrossRef]

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

L. K. Nguyen, M. E. Bravo-Zanoguera, A. L. Kellner, and J. H. Price, “Magnification corrected optical image splitting module for simultaneous multiplanar acquisition,” Proc. SPIE 3921, 31–40 (2000).
[CrossRef]

M. A. Oliva, M. Bravo-Zanoguera, and J. H. Price, “Filtering out contrast reversals for microscopy autofocus,” Appl. Opt. 38, 638–646 (1999).
[CrossRef]

M. Bravo-Zanoguera, B. von Massenbach, A. L. Kellner, and J. H. Price, “High-performance autofocus circuit for biological microscopy,” Rev. Sci. Instrum. 69, 3966–3977 (1998).
[CrossRef]

J. H. Price and D. A. Gough, “Comparison of phase-contrast and fluorescence digital autofocus for scanning microscopy,” Cytometry 16, 283–297 (1994).
[CrossRef] [PubMed]

Prigozhina, N. L.

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

Quercioli, F.

G. Molesini, G. Pedrini, P. Poggi, and F. Quercioli, “Focus-wavelength encoded optical profilometer,” Opt. Commun. 49, 229–233 (1984).
[CrossRef]

Roop, D. R.

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

Shi, K.

Spring, K. R.

S. Inoué and K. R. Spring, Video Microscopy: The Fundamentals, 2nd ed. (Springer, 1997).
[CrossRef]

Steen, W. M.

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Sun, P.-C.

Sun, P.-C. S.

Thieleking, M.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Tiziani, H. J.

Uhde, H.-M.

von Massenbach, B.

M. Bravo-Zanoguera, B. von Massenbach, A. L. Kellner, and J. H. Price, “High-performance autofocus circuit for biological microscopy,” Rev. Sci. Instrum. 69, 3966–3977 (1998).
[CrossRef]

Weinigel, M.

M. Weinigel, A. L. Kellner, and J. H. Price, “Exploration of chromatic aberration for multiplanar imaging: proof of concept with implications for fast, efficient autofocus,” Cytometry A 75A, 999–1006 (2009).
[CrossRef]

Wolf, E.

M. Born and E. Wolf, Principles of Optics (Pergamon, 1980), 6th ed.

Yin, S.

Young, I. T.

F. C. A. Groen, I. T. Young, and G. Ligthart, “A comparison of different focus functions for use in autofocus algorithms,” Cytometry 6, 81–91 (1985).
[CrossRef] [PubMed]

Zacharias, D.

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

Zacharias, D. A.

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

Zatorsky, N. G.

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

Zhong, L.

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

Zhu, L.

Appl. Opt. (5)

Assay Drug Devel. Tech. (3)

P. M. McDonough, R. M. Agustin, R. S. Ingermanson, P. A. Loy, B. M. Buehrer, J. B. Nicoll, N. L. Prigozhina, I. Mikic, and J. H. Price, “Quantification of lipid droplets and associated proteins in cellular models of obesity via high-content/high-throughput microscopy and automated image analysis,” Assay Drug Devel. Tech. 7, 440–460 (2009), PMID: 19895345.
[CrossRef]

M. M. Morelock, E. A. Hunter, T. J. Moran, S. Heynen, C. Laris, M. Thieleking, M. Akong, I. Mikic, S. Callaway, R. P. DeLeon, A. Goodacre, D. Zacharias, and J. H. Price, “Statistics of assay validation in high throughput cell imaging of nuclear factor κB nuclear translocation,” Assay Drug Devel. Tech. 3, 483–499(2005).
[CrossRef]

N. L. Prigozhina, L. Zhong, E. A. Hunter, I. Mikić, S. Callaway, D. R. Roop, M. A. Mancini, D. A. Zacharias, J. H. Price, and P. M. McDonough, “Plasma membrane assays and three-compartment image cytometry for high content screening,” Assay Drug Devel. Tech. 5, 29–48(2007).
[CrossRef]

Cytometry (2)

F. C. A. Groen, I. T. Young, and G. Ligthart, “A comparison of different focus functions for use in autofocus algorithms,” Cytometry 6, 81–91 (1985).
[CrossRef] [PubMed]

J. H. Price and D. A. Gough, “Comparison of phase-contrast and fluorescence digital autofocus for scanning microscopy,” Cytometry 16, 283–297 (1994).
[CrossRef] [PubMed]

Cytometry A (1)

M. Weinigel, A. L. Kellner, and J. H. Price, “Exploration of chromatic aberration for multiplanar imaging: proof of concept with implications for fast, efficient autofocus,” Cytometry A 75A, 999–1006 (2009).
[CrossRef]

J. Biomed. Opt. (1)

M. E. Bravo-Zanoguera, C. A. Laris, L. K. Nguyen, M. Oliva, and J. H. Price, “Dynamic autofocus for continuous-scanning time-delay-and-integration image acquisition in automated microscopy,” J. Biomed. Opt. 12, 034011 (2007).
[CrossRef] [PubMed]

J. Microsc. (1)

Y. Liron, Y. Paran, N. G. Zatorsky, B. Geiger, and Z. Kam, “Laser autofocusing system for high-resolution cell biological imaging,” J. Microsc. 221, 145–151 (2006).
[CrossRef] [PubMed]

Opt. Commun. (1)

G. Molesini, G. Pedrini, P. Poggi, and F. Quercioli, “Focus-wavelength encoded optical profilometer,” Opt. Commun. 49, 229–233 (1984).
[CrossRef]

Opt. Express (1)

Opt. Lasers Eng. (1)

D. P. Hand, M. D. T. Fox, F. M. Haran, C. Peters, S. A. Morgan, M. A. McLean, W. M. Steen, and J. D. C. Jones, “Optical focus control system for laser welding and direct casting,” Opt. Lasers Eng. 34, 415–427 (2000).
[CrossRef]

Opt. Lett. (1)

Proc. SPIE (1)

L. K. Nguyen, M. E. Bravo-Zanoguera, A. L. Kellner, and J. H. Price, “Magnification corrected optical image splitting module for simultaneous multiplanar acquisition,” Proc. SPIE 3921, 31–40 (2000).
[CrossRef]

Rev. Sci. Instrum. (1)

M. Bravo-Zanoguera, B. von Massenbach, A. L. Kellner, and J. H. Price, “High-performance autofocus circuit for biological microscopy,” Rev. Sci. Instrum. 69, 3966–3977 (1998).
[CrossRef]

Scanning (2)

M. A. Browne, O. Akinyemi, and A. Boyde, “Confocal surface profiling utilizing chromatic aberration,” Scanning 14, 145–153 (1992).
[CrossRef]

M. Maly and A. Boyde, “Real-time stereoscopic confocal reflection microscopy using objective lenses with linear longitudinal chromatic dispersion,” Scanning 16, 187–192 (1994).

Other (9)

M. K. Johnson, “Lighting and optical tools for image forensics,” Ph.D. thesis (Dartmouth College, 2007).

Schott, “Optical glass: Data sheets,” http://www.us.schott.com/advanced_optics/english/download/schott_optical_glass_2009_us.pdf (2002).

M. Françon, Progress in Microscopy, Vol.  9 of International Series of Monographs on Pure and Applied Biology(Pergamon, 1961).

S. Inoué and K. R. Spring, Video Microscopy: The Fundamentals, 2nd ed. (Springer, 1997).
[CrossRef]

J. Peters, “Nikon Instruments TiE-PFS dynamic focusing system,” Nature Methods—Application Notes, http://www.nature.com/app_notes/nmeth/2008/082312/full/an6676.html (2008).

P.-C. Cheng, “The contrast formation in optical microscopy,” in Handbook of Biological Confocal Microscopy, 3rd ed. (Springer, 2006), pp. 162–206.
[CrossRef]

L. K. Nguyen, “High-throughput image cytometer for detection of circulating tumor cells and contrast-enhancement filtering for automated 3D image segmentation of cartilage tissue explants,” Ph.D. thesis (University of California–San Diego, 2007).

M. Born and E. Wolf, Principles of Optics (Pergamon, 1980), 6th ed.

H. Bach and N. Neuroth, The Properties of Optical Glass (Springer-Verlag, 1998).

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Figures (6)

Fig. 1
Fig. 1

Longitudinal chromatic aberrations of biconvex BK7 lenses with nominal focal lengths of 25, 37.5, 50, and 75 mm .

Fig. 2
Fig. 2

Schematic of the relay system used to illustrate the control of chromatic aberration generated by the microscope via external optics. The image generated by the microscope at the side port that would normally be imaged onto a camera CCD is reimaged by lenses L 1 and L 2 onto the camera at the new camera image plane.

Fig. 3
Fig. 3

Modulation transfer curves of configuration 2 after the relay optics for various apertures.

Fig. 4
Fig. 4

The measured focal shifts relative to the best focus of 512 nm light for three Nikon 20 × objectives with the Nikon TiE 1 × tube lens are plotted for wavelengths from 412 to 637 nm . The curves fit to the data points are generated using cubic spline interpolation using the MATLAB curve fitting toolbox. The focus was measured in five trials using a 10 μm per division micrometer slide. SDs are not shown, as they are not substantially larger than the data points themselves.

Fig. 5
Fig. 5

The measured focal shifts relative to best focus at 512 nm for three Nikon 20 × objectives [(a)  0.75 NA Plan Apo VC, (b)  0.75 NA Plan Apo, (c)  0.5 NA Plan Fluor] with various lens combinations for the relay optics of Fig. 2 are plotted for wavelengths from 412 to 637 nm . In the legend shown in (c), the first number is the focal length of L 1 in millimeters, the second number is the focal length of L 2 in millimeters, and the last entry is the glass used (FS, fused silica); see also Table 1.

Fig. 6
Fig. 6

Focus function curves for the Nikon 20 × 0.75 NA Plan Apo VC objective used in combination with relay lens configuration 6 of Table 2 ( L 1 and L 2 are 100 mm focal length fused silica lenses) for wavelengths of 412 to 637 nm in 25 nm steps are plotted. These plots illustrate how the wavelength selection can also be used for controlling the spacings of focal planes.

Tables (3)

Tables Icon

Table 1 Coefficients Used for Paraxial Model of Longitudinal Chromatic Aberration a

Tables Icon

Table 2 Relay Lens Combinations Shown in Fig. 1 Used for the Chromatic Aberration Measurements Presented in Fig. 3 a

Tables Icon

Table 3 Performance of Chromatic Aberration Autofocus in Scans of NIH 3T3 Cells on Coverglass a

Equations (10)

Equations on this page are rendered with MathJax. Learn more.

1 f ( λ ) = ( n ( λ ) 1 ) ( C 1 C 2 ) ,
f ( λ ) = f nom ( n nom 1 ) n ( λ ) 1 ,
1 d i ( λ ) = 1 f ( λ ) 1 d o ,
1 d i ( λ ) = n ( λ ) 1 f nom ( n nom 1 ) 1 d o .
n ( λ ) = ( 1 + i = 1 3 B i λ 2 λ 2 C i ) 1 2 ,
W = z z f z α z f z α ,
f z = x y [ h ( x ) i z ( x , y ) ] 2 [ x y i z ( x , y ) ] 2 .
f z = ν = ν 1 ν 2 y w ν f y , z ; ν ,
f y , z ; ν = | x i z ( x , y ) e 2 π j N x ν | 2 ,
f ( z ) = f r ( z ) min ( f r ( z ) ) max ( f r ( z ) ) min ( f r ( z ) ) .

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