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Comparative Observation of the Recombinant Adeno-Associated Virus 2 Using Transmission Electron Microscopy and Atomic Force Microscopy

Published online by Cambridge University Press:  28 September 2007

Heng Chen
Affiliation:
School of Life Science, Shanghai University, Shanghai 200444, P.R. China and College of Life Science & Biotechnology, Shanghai Jiaotong University, Shanghai 200030, P.R. China
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Abstract

Adeno-associated virus (AAV) is a defective, nonpathogenic human parvovirus, which coinfects with a helper adenovirus or herpes virus. AAV's unique characteristics have made it an appealing vector system for gene delivery. AAV or recombinant AAV (rAAV) has been widely detected using negative stain transmission electron microscopy (TEM) but little has been detected using atomic force microscopy (AFM). In this article, we used AFM and TEM to observe the recombinant AAV-2 (rAAV-2) virus particles and applied statistical analysis to the AFM and TEM images. The results indicated that the rAAV-2 particle was a slightly elliptic particle close to round when it was detected by TEM (the mean length of major and minor axes of rAAV-2 particles was 24.77 ± 1.78 nm and 21.84 ± 1.57 nm, respectively), whereas when detected by AFM, the rAAV-2 particle was almost round. Even though the dimensions of the rAAV-2 particle exhibited a polymorphous distribution via off-line particle analysis of AFM, most of the rAAV-2 particles had a mean diameter of approximate 22.04 nm, which was similar to the results obtained by TEM. The results above suggested that AFM was important for accurately determining the average dimensions and distributions of virus particles.

Type
BIOLOGICAL APPLICATIONS
Copyright
© 2007 Microscopy Society of America

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References

REFERENCES

Bartlett, J.S., Wilcher, R. & Samulski, R. (2000). Infectious entry pathway of adeno-associated virus and adeno-associated virus vectors. J Virol 74, 27772785.Google Scholar
Berns, K.I. & Bohenzky, R.A. (1987). Adeno-associated viruses: An update. Adv Virus Res 32, 243306.Google Scholar
Chen, H., Liu, S.P., Chen, L., Huang, J.H. & Xiang, S.M. (2005). Expression of HBcAg mutant with long internal deletion in Saccharomyces cerevisiae and observation of its self-assembly particles by atomic force microscopy (AFM). Int J Biol Macromol 37, 239248.Google Scholar
Chen, H., Lu, J.H., Liang, W.Q., Huang, Y.H., Zhang, W.J. & Zhang, D.B. (2004). Purification and characterization of the recombinant hepatitis B virus core antigen (HBcAg) produced in the yeast Saccharomyces cerevisiae and comparative observation of HBcAg particles by TEM and AFM. Micron 35, 311318.Google Scholar
Crowther, R.A., Kiselev, N.A., Böttcher, B., Berriman, J.A., Borisova, G.P., Ose, V. & Pumpens, P. (1994). Three-dimensional structure of hepatitis B virus core particles determined by electron cryomicroscopy. Cell 77, 943950.Google Scholar
Gao, G.P., Alvira, M.R., Wang, L., Calcedo, R., Johnston, J. & Wilson, J.M. (2002). Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy. Proc Natl Acad Sci USA 99, 1185411859.Google Scholar
Kuznetsov, Y.G., Malkin, A.J., Lucas, R.W., Plomp, M. & McPherson, A. (2001). Imaging of viruses by atomic force microscopy. J Gen Virol 82, 20252034.Google Scholar
Negishi, A., Chen, J.H., McCarty, D.M., Samulski, R.J., Liu, J. & Superfine, R. (2004). Analysis of the interaction between adeno-associated virus and heparan sulfate using atomic force microscopy. Glycobiology 14, 969977.Google Scholar
Ralf, D., Jason, A.K., Stefan, W., Andrea, K. & Jürgen, A.K. (1999). Adeno-associated virus type-2 protein interactions: Formation of pre-encapsidation complexes. J Virol 73, 89898998.Google Scholar
Rigotti, D.J., Kokona, B., Horne, T., Acton, E.K., Lederman, C.D., Johnson, K.A., Manning, R.S., Kane, S.A., Smith, W.F. & Fairman, R. (2005). Quantitative atomic force microscopy image analysis of unusual filaments formed by the canthamoeba castellanii myosin II rod domain. Anal Biochem 346, 189200.Google Scholar
Shi, W. & Bartlett, J.S. (2003). RGD inclusion in VP3 provides adeno-associated virus type 2 (AAV2)-based vectors with a heparan sulfate-independent cell entry mechanism. Am Soc Gene Ther Mol Ther 7, 515525.Google Scholar
Srivastava, A., Lusby, E.W. & Berns, K.I. (1983). Nucleotide sequence and organization of the adeno-associated virus 2 genome. J Virol 45, 555564.Google Scholar
Timpe, J., Bevington, J., Casper, J., Dignam, D. & Trempe, J.P. (2005). Mechanisms of adeno-associated virus genome encapsidation. Curr Gene Ther 5, 273284.Google Scholar
Wright, J.F., Le, T., Prado, J., Davidson, J.B., Smith, P.H., Zhen, Z., Sommer, J.M., Pierce, G.F. & Qu, G. (2005). Identification of factors that contribute to recombinant AAV2 particle aggregation and methods to prevent its occurrence during vector purification and formulation. Mol Ther 12, 171178.Google Scholar
Xie, Q., Hare, J., Turnigan, J. & Chapman, M.S. (2004). Large-scale production, purification and crystallization of wild-type adeno-associated virus-2. J Virol Methods 122, 1727.Google Scholar
Yamaguchi, M., Hirano, T., Hirokawa, H., Sugahara, K., Mizokami, H. & Matsubara, K. (1988). Cryo-electron microscopy of hepatitis B virus core particles produced by transformed yeast: Comparison with negative staining and ultrathin sectioning. J Electron Microsc 37, 337341.Google Scholar