Abstract

We report a fast fluorescence imaging flow cytometer for phytoplankton analysis that can achieve a volume flow rate up to 1ml/min. The instrument shows a high immunity to motion blur in image captured with a lateral resolution of 0.75 ± 0.06 μm for a wide size range ~1 μm to ~200 μm. This is made possible by suppressing the out-of-focus light using thin light sheet illumination and image deconvolution, and by precluding the motion-blur with a unique flow configuration. Preliminary results from untreated coastal water samples show the technique has high potential as a practical field instrument for monitoring phytoplankton abundance and species composition.

© 2013 Optical Society of America

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References

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  1. Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
    [CrossRef]
  2. J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
    [CrossRef] [PubMed]
  3. M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
    [PubMed]
  4. J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
    [CrossRef] [PubMed]
  5. V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
    [CrossRef] [PubMed]
  6. V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).
  7. K. Goda, K. K. Tsia, and B. Jalali, “Serial time-encoded amplified imaging for real-time observation of fast dynamic phenomena,” Nature458(7242), 1145–1149 (2009).
    [CrossRef] [PubMed]
  8. D. A. Basiji and W. E. Ortyn, “Imaging and analyzing parameters of small moving objects such as cells,” Amnis Corporation, assignee. US Patent 6211955, 2000–03–29 (2001).
  9. S. S. Gorthi, D. Schaak, and E. Schonbrun, “Fluorescence imaging of flowing cells using a temporally coded excitation,” Opt. Express21(4), 5164–5170 (2013).
    [CrossRef] [PubMed]
  10. R. J. Olson and H. M. Sosik, “A submersible imaging-in-flow instrument to analyze nano and microplankton: Imaging FlowCytobot,” Limnol. Oceanogr. Methods5, 195–203 (2007).
    [CrossRef]
  11. W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
    [CrossRef] [PubMed]
  12. C. K. Sieracki, M. E. Sieracki, and C. S. Yentsch, “An imaging-in-flow system for automated analysis of marine microplankton,” Mar. Ecol. Prog. Ser.168, 285–296 (1998).
    [CrossRef]
  13. V. Kachel and J. Wietzorrek, “Flow cytometry and integrated imaging,” Sci. Mar.64(2), 247–254 (2000).
  14. B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
    [CrossRef] [PubMed]
  15. J. Wu, J. Li, and R. K. Y. Chan, “A light sheet based high throughput 3D-imaging flow cytometer for phytoplankton analysis,” Opt. Express21(12), 14474–14480 (2013).
    [CrossRef] [PubMed]
  16. K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope,” Rev. Sci. Instrum.78(2), 023705 (2007).
    [CrossRef] [PubMed]
  17. E. Fuchs, J. S. Jaffe, R. A. Long, and F. Azam, “Thin laser light sheet microscope for microbial oceanography,” Opt. Express10(2), 145–154 (2002).
    [CrossRef] [PubMed]
  18. J. A. Conchello and M. E. Dresser, “Extended depth-of-focus microscopy via constrained deconvolution,” J. Biomed. Opt.12(6), 064026 (2007).
    [CrossRef] [PubMed]
  19. Hong Kong Environmental Protection Department, “Marine Water Quality Reports,” (2013). http://www.epd.gov.hk/epd/english/environmentinhk/water/marine_quality/mwq_report.html .

2013 (2)

2012 (1)

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
[CrossRef] [PubMed]

2011 (1)

B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
[CrossRef] [PubMed]

2010 (1)

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

2009 (1)

K. Goda, K. K. Tsia, and B. Jalali, “Serial time-encoded amplified imaging for real-time observation of fast dynamic phenomena,” Nature458(7242), 1145–1149 (2009).
[CrossRef] [PubMed]

2007 (4)

R. J. Olson and H. M. Sosik, “A submersible imaging-in-flow instrument to analyze nano and microplankton: Imaging FlowCytobot,” Limnol. Oceanogr. Methods5, 195–203 (2007).
[CrossRef]

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope,” Rev. Sci. Instrum.78(2), 023705 (2007).
[CrossRef] [PubMed]

J. A. Conchello and M. E. Dresser, “Extended depth-of-focus microscopy via constrained deconvolution,” J. Biomed. Opt.12(6), 064026 (2007).
[CrossRef] [PubMed]

2002 (1)

2000 (1)

V. Kachel and J. Wietzorrek, “Flow cytometry and integrated imaging,” Sci. Mar.64(2), 247–254 (2000).

1999 (1)

M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
[PubMed]

1998 (1)

C. K. Sieracki, M. E. Sieracki, and C. S. Yentsch, “An imaging-in-flow system for automated analysis of marine microplankton,” Mar. Ecol. Prog. Ser.168, 285–296 (1998).
[CrossRef]

1989 (1)

J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
[CrossRef] [PubMed]

1980 (1)

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

1979 (1)

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

Ahrens, O.

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

Azam, F.

Basiji, D. A.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Beardall, J.

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Benker, G.

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

Boddy, L.

M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
[PubMed]

Boulanger, C. M.

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
[CrossRef] [PubMed]

Chan, R. K. Y.

Cheung, M. C.

B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
[CrossRef] [PubMed]

Conchello, J. A.

J. A. Conchello and M. E. Dresser, “Extended depth-of-focus microscopy via constrained deconvolution,” J. Biomed. Opt.12(6), 064026 (2007).
[CrossRef] [PubMed]

Dresser, M. E.

J. A. Conchello and M. E. Dresser, “Extended depth-of-focus microscopy via constrained deconvolution,” J. Biomed. Opt.12(6), 064026 (2007).
[CrossRef] [PubMed]

Dubelaar, G. B. J.

J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
[CrossRef] [PubMed]

Ehrlich, D. J.

B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
[CrossRef] [PubMed]

Evans, J. G.

B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
[CrossRef] [PubMed]

Finkel, Z. V.

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Flynn, K. J.

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Frost, K.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Fuchs, E.

Glossner, E.

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

Goda, K.

K. Goda, K. K. Tsia, and B. Jalali, “Serial time-encoded amplified imaging for real-time observation of fast dynamic phenomena,” Nature458(7242), 1145–1149 (2009).
[CrossRef] [PubMed]

Gorthi, S. S.

Greger, K.

K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope,” Rev. Sci. Instrum.78(2), 023705 (2007).
[CrossRef] [PubMed]

Guerin, C. L.

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
[CrossRef] [PubMed]

Hall, B. E.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Jaffe, J. S.

Jalali, B.

K. Goda, K. K. Tsia, and B. Jalali, “Serial time-encoded amplified imaging for real-time observation of fast dynamic phenomena,” Nature458(7242), 1145–1149 (2009).
[CrossRef] [PubMed]

Jonker, R. R.

M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
[PubMed]

Kachel, V.

V. Kachel and J. Wietzorrek, “Flow cytometry and integrated imaging,” Sci. Mar.64(2), 247–254 (2000).

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

Le Van Kim, C.

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
[CrossRef] [PubMed]

Li, J.

Liang, L.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Lichtnau, K.

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

Long, R. A.

McKenna, B. K.

B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
[CrossRef] [PubMed]

Morris, C. W.

M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
[PubMed]

Olson, R. J.

R. J. Olson and H. M. Sosik, “A submersible imaging-in-flow instrument to analyze nano and microplankton: Imaging FlowCytobot,” Limnol. Oceanogr. Methods5, 195–203 (2007).
[CrossRef]

Ortyn, W. E.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Peeters, J. C. H.

J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
[CrossRef] [PubMed]

Perry, D. J.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Picot, J.

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
[CrossRef] [PubMed]

Quigg, A.

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Raven, J. A.

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Rees, T. A. V.

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Ringelberg, J.

J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
[CrossRef] [PubMed]

Schaak, D.

Schonbrun, E.

Sieracki, C. K.

C. K. Sieracki, M. E. Sieracki, and C. S. Yentsch, “An imaging-in-flow system for automated analysis of marine microplankton,” Mar. Ecol. Prog. Ser.168, 285–296 (1998).
[CrossRef]

Sieracki, M. E.

C. K. Sieracki, M. E. Sieracki, and C. S. Yentsch, “An imaging-in-flow system for automated analysis of marine microplankton,” Mar. Ecol. Prog. Ser.168, 285–296 (1998).
[CrossRef]

Sosik, H. M.

R. J. Olson and H. M. Sosik, “A submersible imaging-in-flow instrument to analyze nano and microplankton: Imaging FlowCytobot,” Limnol. Oceanogr. Methods5, 195–203 (2007).
[CrossRef]

Stelzer, E. H. K.

K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope,” Rev. Sci. Instrum.78(2), 023705 (2007).
[CrossRef] [PubMed]

Swoger, J.

K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope,” Rev. Sci. Instrum.78(2), 023705 (2007).
[CrossRef] [PubMed]

Tsia, K. K.

K. Goda, K. K. Tsia, and B. Jalali, “Serial time-encoded amplified imaging for real-time observation of fast dynamic phenomena,” Nature458(7242), 1145–1149 (2009).
[CrossRef] [PubMed]

Valet, G.

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

Venkatachalam, V.

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Visser, J. W. M.

J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
[CrossRef] [PubMed]

Weiss, W.

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

Wietzorrek, J.

V. Kachel and J. Wietzorrek, “Flow cytometry and integrated imaging,” Sci. Mar.64(2), 247–254 (2000).

Wilkins, M. F.

M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
[PubMed]

Wu, J.

Yentsch, C. S.

C. K. Sieracki, M. E. Sieracki, and C. S. Yentsch, “An imaging-in-flow system for automated analysis of marine microplankton,” Mar. Ecol. Prog. Ser.168, 285–296 (1998).
[CrossRef]

Appl. Environ. Microbiol. (1)

M. F. Wilkins, L. Boddy, C. W. Morris, and R. R. Jonker, “Identification of phytoplankton from flow cytometry data by using radial basis function neural networks,” Appl. Environ. Microbiol.65(10), 4404–4410 (1999).
[PubMed]

Cytometry (1)

J. C. H. Peeters, G. B. J. Dubelaar, J. Ringelberg, and J. W. M. Visser, “Optical plankton analyser: a flow cytometer for plankton analysis, I: design considerations,” Cytometry10(5), 522–528 (1989).
[CrossRef] [PubMed]

Cytometry A (1)

W. E. Ortyn, D. J. Perry, V. Venkatachalam, L. Liang, B. E. Hall, K. Frost, and D. A. Basiji, “Extended depth of field Imaging for high speed cell analysis,” Cytometry A71(4), 215–231 (2007).
[CrossRef] [PubMed]

Cytotechnology (1)

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology64(2), 109–130 (2012).
[CrossRef] [PubMed]

Flow Cytometry (1)

V. Kachel, G. Benker, W. Weiss, E. Glossner, G. Valet, and O. Ahrens, “Problems of fast imaging in flow,” Flow CytometryIV, 45–49 (1980).

J. Biomed. Opt. (1)

J. A. Conchello and M. E. Dresser, “Extended depth-of-focus microscopy via constrained deconvolution,” J. Biomed. Opt.12(6), 064026 (2007).
[CrossRef] [PubMed]

J. Histochem. Cytochem. (1)

V. Kachel, G. Benker, K. Lichtnau, G. Valet, and E. Glossner, “Fast imaging in flow: a means of combining flow-cytometry and image analysis,” J. Histochem. Cytochem.27(1), 335–341 (1979).
[CrossRef] [PubMed]

J. Plankton Res. (1)

Z. V. Finkel, J. Beardall, K. J. Flynn, A. Quigg, T. A. V. Rees, and J. A. Raven, “Phytoplankton in a changing world: cell size and elemental stoichiometry,” J. Plankton Res.32(1), 119–137 (2010).
[CrossRef]

Limnol. Oceanogr. Methods (1)

R. J. Olson and H. M. Sosik, “A submersible imaging-in-flow instrument to analyze nano and microplankton: Imaging FlowCytobot,” Limnol. Oceanogr. Methods5, 195–203 (2007).
[CrossRef]

Mar. Ecol. Prog. Ser. (1)

C. K. Sieracki, M. E. Sieracki, and C. S. Yentsch, “An imaging-in-flow system for automated analysis of marine microplankton,” Mar. Ecol. Prog. Ser.168, 285–296 (1998).
[CrossRef]

Nat. Methods (1)

B. K. McKenna, J. G. Evans, M. C. Cheung, and D. J. Ehrlich, “A parallel microfluidic flow cytometer for high-content screening,” Nat. Methods8(5), 401–403 (2011).
[CrossRef] [PubMed]

Nature (1)

K. Goda, K. K. Tsia, and B. Jalali, “Serial time-encoded amplified imaging for real-time observation of fast dynamic phenomena,” Nature458(7242), 1145–1149 (2009).
[CrossRef] [PubMed]

Opt. Express (3)

Rev. Sci. Instrum. (1)

K. Greger, J. Swoger, and E. H. K. Stelzer, “Basic building units and properties of a fluorescence single plane illumination microscope,” Rev. Sci. Instrum.78(2), 023705 (2007).
[CrossRef] [PubMed]

Sci. Mar. (1)

V. Kachel and J. Wietzorrek, “Flow cytometry and integrated imaging,” Sci. Mar.64(2), 247–254 (2000).

Other (2)

Hong Kong Environmental Protection Department, “Marine Water Quality Reports,” (2013). http://www.epd.gov.hk/epd/english/environmentinhk/water/marine_quality/mwq_report.html .

D. A. Basiji and W. E. Ortyn, “Imaging and analyzing parameters of small moving objects such as cells,” Amnis Corporation, assignee. US Patent 6211955, 2000–03–29 (2001).

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Figures (5)

Fig. 1
Fig. 1

Schematic diagram of the 2D fluorescence imaging flow cytometer. (a) side view; (b) top view.

Fig. 2
Fig. 2

Experimentally measured lateral PSF. (a) Integrated-intensity projection of the 3D PSF of the imaging optics, 51 × 51pixels; (b) Intensity profile of the PSF, FWHM is 0.75 ± 0.06 μm.

Fig. 3
Fig. 3

Images obtained from natural coastal water samples. (a) Single frame; (b) Maximum projection of 60 deconvolved images. Experimental conditions: volume rate is 1 ml/min; exposure time is 1 s. Scale bars: 20 μm.

Fig. 4
Fig. 4

Comparison between original and deconvolved image. (a) Original image; (b) Deconvolved image. White curves are the intensity profiles along the red lines selected. Scale bars: 10 μm.

Fig. 5
Fig. 5

Imagery of phytoplankton with different morphologies obtained from natural coastal water samples.

Equations (1)

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G( x i , y i )=H( x i , y i , x o , y o )S( x o , y o ).

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