Skip to main content
SpringerLink
Log in

Automated tracking and analysis of phospholipid vesicle contours in phase contrast microscopy images

  • Original Article
  • Published:
Medical & Biological Engineering & Computing Aims and scope Submit manuscript

Abstract

In this article, we propose a method for automated tracking and analysis of vesicle contours in video sequences acquired by phase contrast microscopy. The contour is determined in each frame of the selected video sequence by detecting the transition between the interior and exterior of the vesicle that is reflected in the image intensity gradients. The resulting contour points are represented in the polar coordinate system, i.e., with uniform angular sampling and with coordinates that originate from the vesicle center of mass, enabling the analysis of the vesicle shape and its membrane fluctuations. By analyzing artificial images with known ground-truth contours, the accuracy and precision of the proposed method was estimated to be 34.1 and 26.9 nm for image signal-to-noise ratio of 23 dB and pixel size of 35 nm, respectively. The proposed method was evaluated on quasi-spherical vesicles made up of different proportions of 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-Phosphocholine (POPC) and cholesterol and exposed to different temperatures. The results show that the method is robust and efficient in terms of speed and quantitative description of vesicle fluctuations. The magnitude of vesicle membrane fluctuations increased with temperature, while the bending rigidity of the membrane was increasing for temperatures up to 20°C and decreasing for higher temperatures irrespective of the vesicle molecular structure.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  1. Ambriz-Colin F, Torres-Cisneros M, Avina-Cervantes J, Saavedra-Martinez J, Debeir O, Sanchez-Mondragon J (2006) Detection of biological cells in phase-contrast microscopy images. In: Gelbukh A, Reyes C (eds) Proceedings of the fifth mexican international conference on artificial intelligence, IEEE Computer Society Washington, USA, pp 68–77

  2. Angelova M, Soléau S, Méléard P, Faucon F, Bothorel P (1992) Preparation of giant vesicles by external AC electric fields. Kinetics and applications. In: Helm C, Lösche M, Möhwald H (eds) Trends in colloid and interface science VI, progress in colloid and polymer science. Springer, Berlin, pp 127–131

    Chapter  Google Scholar 

  3. Bitler A, Barbul A, Korenstein R (1999) Detection of movement at the erythrocyte’s edge by scanning phase contrast microscopy. J Microsc 193:171–178

    Article  PubMed  CAS  Google Scholar 

  4. Bivas I, Hanusse P, Bothorel P, Lalanne J, Aguerre-Chariol O (1987) An application of the optical microscopy to the determination of the curvature elastic modulus of biological and model membranes. J Phys (Paris) 48:855–867

    Google Scholar 

  5. Brochard F, Lennon J (1975) Frequency spectrum of the flicker phenomenon in erythrocytes. J Phys (Paris) 36:1035–1047

    Article  Google Scholar 

  6. Bunyak F, Palaniappan K, Nath SK, Baskin TI, Dong G (2006) Quantitative cell motility for in vitro wound healing using level set-based active contour tracking. In: Kovačević J, Meijering E (eds) Proceedings of IEEE international symposium on biomedical imaging: from nano to macro, IEEE Service Center, Piscataway, NJ, pp 1040–1043

  7. Debeir O, Van Ham P, Kiss R, Decaestecker C (2005) Tracking of migrating cells under phase-contrast video microscopy with combined mean-shift processes. IEEE Trans Med Imaging 24:697–711

    Article  PubMed  CAS  Google Scholar 

  8. Döbereiner HG (2000) Properties of giant vesicles. Curr Opin Colloid Interface Sci 5:256–263

    Article  Google Scholar 

  9. Döbereiner HG, Evans E, Kraus M, Seifert U, Wortis M (1997) Mapping vesicle shapes into the phase diagram: a comparison of experiment and theory. Phys Rev E Stat Phys Plasmas Fluids Relat Interdisciplin Top 55:4458–4474

    Google Scholar 

  10. Döbereiner HG, Gompper G, Haluska C, Kroll D, Petrov P, Riske K (2003) Advanced flicker spectroscopy of fluid membranes. Phys Rev Lett 91:048301

    Article  PubMed  Google Scholar 

  11. Duwe H, Kaes J, Sackmann E (1990) Bending elastic moduli of lipid bilayers: modulation by solutes. J Phys (Paris) 51:945–961

    CAS  Google Scholar 

  12. Engelhardt H, Duwe H, Sackmann E (1985) Bilayer bending elasticity measured by Fourier analysis of thermally excited surface undulations of flaccid vesicles. J Phys Lett (Paris) 46:395–400

    CAS  Google Scholar 

  13. Ersoy I, Bunyak F, Palaniappan K, Sun M, Forgacs G (2008) Cell spreading analysis with directed edge profile-guided level set active contours. In: Metaxas D, Axel L, Fichtinger G, Székely G (eds) Proceedings of the medical image computing and computer-assisted intervention, Springer, Berlin, pp 376–383

  14. Evans J, Gratzer W, Mohandas N, Parker K, Sleep J (2008) Fluctuations of the red blood cell membrane: relation to mechanical properties and lack of ATP dependence. Biophys J 94:4134–4144

    Article  PubMed  CAS  Google Scholar 

  15. Faucon J, Mitov MD, Méléard P, Bivas I, Bothorel P (1989) Bending elasticity and thermal fluctuations of lipid membranes. Theoretical and experimental requirements. J Phys (Paris) 50:2389–2414

    Google Scholar 

  16. Hägerstrand H, Danieluk M, Bobrowska-Hägerstrand M, Pector V, Ruysschaert J, Kralj-Iglic V, Iglic A (1999) Liposomes composed of a double-chain cationic amphiphile (Vectamidine) induce their own encapsulation into human erythrocytes. Biochimica et Biophysica Acta (BBA)—Biomembranes 1421:125–130

    Article  Google Scholar 

  17. Immerkær J (1996) Fast noise variance estimation. Comput Vis Image Underst 64:300–302

    Article  Google Scholar 

  18. Jones MM, Chapman D (1995) Micelles, monolayers and biomembranes. Wiley-Liss, New York

    Google Scholar 

  19. Kralj-Iglič V, Gomišček G, Majhenc J, Arrigler V, Svetina S (2001) Myelin-like protrusions of giant phospholipid vesicles prepared by electroformation. Colloids Surf A 181:315–318

    Article  Google Scholar 

  20. Leekumjorn S, Sum AK (2007) Molecular characterization of gel and liquid-crystalline structures of fully hydrated POPC and POPE bilayers. J Phys Chem B 111:6026–6033

    Article  PubMed  CAS  Google Scholar 

  21. Marsh D (2006) Elastic curvature constants of lipid monolayers and bilayers. Chem Phys Lipids 144:146–159

    Article  PubMed  CAS  Google Scholar 

  22. Méléard P, Gerbeaud C, Pott T, Fernandez-Puente L, Bivas I, Mitov M, Dufourcq J, Bothorel P (1997) Bending elasticities of model membranes: influences of temperature and sterol content. Biophys J 72:2616–2629

    Article  PubMed  Google Scholar 

  23. Milner ST, Safran SA (1987) Dynamical fluctuations of droplet microemulsions and vesicles. Phys Rev A 36:4371

    Article  PubMed  CAS  Google Scholar 

  24. Mukherjee D, Ray N, Acton S (2004) Level set analysis for leukocyte detection and tracking. IEEE Trans Image Process 13:562–572

    Article  PubMed  Google Scholar 

  25. Mutz M, Helfrich W (1990) Bending rigidities of some biological model membranes as obtained from the Fourier analysis of contour sections. J Phys (Paris) 51:991–1001

    CAS  Google Scholar 

  26. Niggemann G, Kummrow M, Helfrich W (1995) The bending rigidity of phosphatidylcholine bilayers: dependences on experimental method, sample cell sealing and temperature. J Phys II 5:413–425

    Article  CAS  Google Scholar 

  27. Nikolov V, Lipowsky R, Dimova R (2007) Behavior of giant vesicles with anchored DNA molecules

  28. Pécréaux J, Döbereiner H, Prost J, Joanny J, Bassereau P (2004) Refined contour analysis of giant unilamellar vesicles. Eur Phys J E 13:277–290

    Article  PubMed  Google Scholar 

  29. Sawant RR, Torchilin VP (2010) Liposomes as ‘smart’ pharmaceutical nanocarriers. Soft Matter 6:4026

    Article  CAS  Google Scholar 

  30. Schneider M, Jenkins J, Webb W (1984) Thermal fluctuations of large quasi-spherical bimolecular phospholipid vesicles. J Phys (Paris) 45:1457–1472

    CAS  Google Scholar 

  31. Sheppard J, Wilson T (1981) The halo effect of image processing by spatial frequency filtering. Optik 56:19–23

    Google Scholar 

  32. Uzoigwe C (2006) The human erythrocyte has developed the biconcave disc shape to optimise the flow properties of the blood in the large vessels. Med Hypotheses 67:1159–1163

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

This study has been supported by the Ministry of Higher Education, Science and Technology, Slovenia, under grants P2-0232, L2-7381, L2-9758, L2-2023, J2-0716, and J7-2246. The authors also thank A. Iglič and J. Pavlič from the University of Ljubljana, Faculty of Electrical Engineering, for their assistance in specimen preparation and image acquisition, and M. Frank from the University of Ljubljana, Faculty of Medicine, for providing images of erythrocytes.

Author information

Authors and Affiliations

  1. Faculty of Electrical Engineering, University of Ljubljana, Ljubljana, Slovenia

    Peter Usenik, Tomaž Vrtovec, Franjo Pernuš & Boštjan Likar

Authors
  1. Peter Usenik
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tomaž Vrtovec
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Franjo Pernuš
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Boštjan Likar
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Peter Usenik.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Usenik, P., Vrtovec, T., Pernuš, F. et al. Automated tracking and analysis of phospholipid vesicle contours in phase contrast microscopy images. Med Biol Eng Comput 49, 957–966 (2011). https://doi.org/10.1007/s11517-011-0789-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11517-011-0789-0

Keywords

Search

Navigation