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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  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,” Cytometry 10(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,” Cytotechnology 64(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 Cytometry IV, 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,” Nature 458(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. Express 21(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. Methods 5, 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 A 71(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. Methods 8(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. Express 21(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. Express 10(2), 145–154 (2002).
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  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]
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2013 (2)

2012 (1)

J. Picot, C. L. Guerin, C. Le Van Kim, and C. M. Boulanger, “Flow cytometry: retrospective, fundamentals and recent instrumentation,” Cytotechnology 64(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. Methods 8(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,” Nature 458(7242), 1145–1149 (2009).
[Crossref] [PubMed]

2007 (4)

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

R. J. Olson and H. M. Sosik, “A submersible imaging-in-flow instrument to analyze nano and microplankton: Imaging FlowCytobot,” Limnol. Oceanogr. Methods 5, 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 A 71(4), 215–231 (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,” Cytometry 10(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 Cytometry IV, 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 Cytometry IV, 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 A 71(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 Cytometry IV, 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,” Cytotechnology 64(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. Methods 8(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,” Cytometry 10(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. Methods 8(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. Methods 8(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 A 71(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 Cytometry IV, 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,” Nature 458(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,” Cytotechnology 64(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 A 71(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,” Nature 458(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 Cytometry IV, 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,” Cytotechnology 64(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 A 71(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. Methods 8(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. Methods 5, 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 A 71(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,” Cytometry 10(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 A 71(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,” Cytotechnology 64(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,” Cytometry 10(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. Methods 5, 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,” Nature 458(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 Cytometry IV, 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 A 71(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,” Cytometry 10(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 Cytometry IV, 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,” Cytometry 10(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 A 71(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,” Cytotechnology 64(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 Cytometry IV, 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. Methods 5, 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. Methods 8(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,” Nature 458(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.

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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.

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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.

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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.

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Fig. 5
Fig. 5

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

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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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