Advertisement

Liposomes pp 319-331 | Cite as

Environmental Scanning Electron Microscope Imaging of Vesicle Systems

  • Yvonne PerrieEmail author
  • Habib Ali
  • Daniel J. Kirby
  • Afzal U. R. Mohammed
  • Sarah E. McNeil
  • Anil Vangala
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 606)

Abstract

The structural characteristics of liposomes have been widely investigated and there is certainly a strong understanding of their morphological characteristics. Imaging of these systems, using techniques such as freeze-fracturing methods, transmission electron microscopy, and cryo-electron imaging, has allowed us to appreciate their bilayer structures and factors that influence this. However, there are a few methods that study these systems in their natural hydrated state; commonly, the liposomes are visualized after drying, staining and/or fixation of the vesicles. Environmental scanning electron microscopy (ESEM) offers the ability to image a liposome in its hydrated state without the need for prior sample preparation. We were the first to use ESEM to study the liposomes and niosomes, and have been able to dynamically follow the hydration of lipid films and changes in liposome suspensions as water condenses onto, or evaporates from, the sample in real-time. This provides an insight into the resistance of liposomes to coalescence during dehydration, thereby providing an alternative assay for liposome formulation and stability.

Key words

Liposomes Surfactant vesicles Niosomes Non-ionic surfactant vesicles Lipoplexes ESEM analysis 

Notes

Acknowledgments

The authors gratefully acknowledge the financial support rendered: during the time of this work, Afzal Mohammed’s research was supported by Pfizer Global Research and The School of Life and Health Sciences, Aston University; Anil Vangala was awarded an Aston Scholarship; Daniel Kirby was funded by the European Commission (contract no. LSHP-CT-2003-503367), Habib Ali and Sarah McNeil were both funded through EPSRC Case awards, with additional support from Lipoxen Technologies Ltd for Sarah McNeil.

References

  1. 1.
    Segal AW, Willis EJ, Richmond JE, Slavin G, Black CD, Gregoriadis G (1974) Morpho­logicalobservations on the cellular and subcellular destination of intravenously administered liposomes. Br J Exp Pathol 55:320-7PubMedGoogle Scholar
  2. 2.
    Sternberg B, Sorgi FL, Huang L (1994) New structures in complex formation between DNA and cationic liposomes visualized by freeze—fracture electron microscopy. FEBS Lett 356:361-6CrossRefPubMedGoogle Scholar
  3. 3.
    Xu Y, Hui S-W, Frederik P, Szoka FC Jr (1999) Pysicochemical characterisation and purification of cationic lipoplexes. Biophysical J 77:341-53CrossRefGoogle Scholar
  4. 4.
    Perrie Y, Frederik PM, Gregoriadis G (2001) Liposome-mediated immunisation: the effect of vesicle composition. Vaccine 19:3301-10CrossRefPubMedGoogle Scholar
  5. 5.
    Davidsen J, Rosenkrands I, Christensen D, Vangala A, Kirby D, Perrie Y, Agger EM, Andersen P (2005) Characterization of cationic liposomes based on dimethyldioctadecylammonium and synthetic cord factor from M. tuberculosis (trehalose 6, 6′-dibehenate) - a novel adjuvant inducing both strong CMI and antibody responses. Biochim Biophys Acta 1718(1-2):22-31CrossRefPubMedGoogle Scholar
  6. 6.
    Arunothayanun P, Turton JA, Uchegbu IF, Florence AT (1999) Preparation and in vitro/in vivo evaluation of luteinizing hormone releasing hormone (LHRH)-loaded polyhedral and spherical/tubular niosomes. J Pharm Sci 88:34-8CrossRefPubMedGoogle Scholar
  7. 7.
    Mohammed AR, Weston N, Coombes AGA, Fitzgerald M, Perrie Y (2004) Liposome formulation of poorly water soluble drugs: optimisation of drug loading and ESEM analysis of stability. Int J Pharm 285:23-34CrossRefPubMedGoogle Scholar
  8. 8.
    Donald AM (1998) Taking SEMs into a new environment. Mater World: 399-401Google Scholar
  9. 9.
    Egelhaaf RM, Epand RF, Maekawa S (2003) The arrangement of cholesterol in membranes and binding of NAP-22. Chem Phys Lipids 122:33-9CrossRefGoogle Scholar
  10. 10.
    McKinlay KJ, Allison FJ, Scotchford CA, Grant DM, Oliver JM, King JR, Wood JV, Brown PD (2004) Comparison of environmental scanning electron microscopy with high vacuum scanning electron microscopy as applied to the assessment of cell morphology. J Biomed Mater Res 69A:359-66CrossRefGoogle Scholar
  11. 11.
    Muscariello L, Rosso F, Marino G, Giordano A, Barbarisi M, Cafiero G, Barbarisi A (2005) A critical overview of ESEM applications in the biological field. J Cell Physiol 205:328-34CrossRefPubMedGoogle Scholar
  12. 12.
    Robinson VN (1975) A wet stage modification to a scanning electron microscope. J Microsc 103:71-7PubMedGoogle Scholar
  13. 13.
    Moncrieff DA, Robinson VNE, Harris LB (1978) Charge neutralisation of insulating surfaces in the SEM by gas ionisation. J Phys D: Appl Phys 11:2315-25CrossRefGoogle Scholar
  14. 14.
    Moncrieff DA, Barker PR, Robinson VNE (1979) Electron scattering by gas in the scanning electron microscope. J Phys D: Appl Phys 12:481-8CrossRefGoogle Scholar
  15. 15.
    Danilatos GD (1993) Introduction to the ESEM instrument. Microsc Res Tech 25:354-61CrossRefPubMedGoogle Scholar
  16. 16.
    Danilatos GD (1990) Theory of the gaseous detector device in the ESEM. Adv Elect Electron Phy 78:1-102Google Scholar
  17. 17.
    Danilatos GD (1988) Foundations of environmental scanning electron microscope. Adv Elect Electron Phy 71:109-250Google Scholar
  18. 18.
    Donald AM, He C, Royall CP, Sferrazza M, Stelmashenko NA, Thiel BL (2000) Applications of environmental scanning electron microscopy to colloidal aggregation and film formation. Colloid Surface Physicochem Eng Aspect 174:37-53CrossRefGoogle Scholar
  19. 19.
    Stokes DJ (2003) Recent advances in electron imaging, image interpretation and applications: environmental scanning electron microscopy. Philos Transact A Math Phys Eng Sci 361:2771-87CrossRefPubMedGoogle Scholar
  20. 20.
    Perrie Y, Mohammed AR, Vangala A, McNeil SE (2007) Environmental scanning electron microscopy studies of liposomes and niosomes. J Liposome Res 17(1):27-37CrossRefPubMedGoogle Scholar
  21. 21.
    Vangala AK, Kirby D, Rosenkrands I, Agger E-M, Andersen P, Perrie Y (2006) A comparative study of cationic liposomes and niosome-based adjuvant systems for protein subunit vaccines Characterisation, Environmental Scan­ning Electron Microscopy analysis and immunisation studies. J Pharm Pharmacol 58:787-99CrossRefPubMedGoogle Scholar
  22. 22.
    Elvira C, Fanovich A, Fernandez M, Fraile J, San RJ, Domingo C (2004) Evaluation of drug delivery characteristics of microspheres of PMMA-PCL-cholesterol obtained by supercritical-CO2 impregnation and by dissolution-evaporation techniques. J Control Release 99:231-40CrossRefPubMedGoogle Scholar
  23. 23.
    D’Emanuele A, Dinarvand R (1995) Preparation, characterisation, and drug release from thermoresponsive microspheres. Int J Pharm 118:237-42CrossRefGoogle Scholar
  24. 24.
    Cao Y, Li H (2002) Interfacial activity of a novel family of polymeric surfactants. Eur Polymer J 38:1457-63CrossRefGoogle Scholar
  25. 25.
    Bangham AD, Standish MM, Watkins JC (1965) Diffusion of univalent ions across the lamellae of swollen phospholipids. J Mol Biol 13:328-52Google Scholar
  26. 26.
    Kirby C, Gregoriadis G (1984) Dehydration-rehydration vesicles: a simple method for high yield drug entrapment in liposomes. Biotechnology 2:979-84CrossRefGoogle Scholar

Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Yvonne Perrie
    • 1
    Email author
  • Habib Ali
    • 1
  • Daniel J. Kirby
    • 1
  • Afzal U. R. Mohammed
    • 1
  • Sarah E. McNeil
    • 1
  • Anil Vangala
    • 2
  1. 1.School of Life and Health SciencesAston UniversityBirminghamUK
  2. 2.School of Pharmacy and ChemistryKingston UniversityLondonUK

Personalised recommendations