Evaluation of Immunoassays for Electron Microscopy

  • Frances W. Doane
  • Nan Anderson
  • Francis Lee
  • Kathryn Pegg-Feige
  • John Hopley


In recent years immunoelectron microscopy (IEM) has become one of the several immunoassays available to virologists for detection and identification of viruses. Like other immunological assays, the reliability depends on careful attention to test conditions (e.g. antigen/antibody concentration, incubation time, temperature, pH) and to a regard for incorporation of a range of appropriate controls. This chapter will discuss several IEM techniques that can be applied to fluid specimens commonly encountered in diagnostic virology. Consideration will be given to some of the variables that affect the efficiency of these tests, and to the advantages and disadvantages of IEM relative to other immunoassays. Additional practical details are available from several published sources (Doane, 1974, 1986, 1987, 1988a, 1988b; Doane and Anderson, 1977, 1987; Katz and Kohn, 1984; Kjeldsberg, 1986).


Virus Particle Tobacco Mosaic Virus Immunoelectron Microscopy Rubella Virus Agar Diffusion Method 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Almeida J, Cinader B, Howatson A (1963) The structure of antigen-antibody complexes. A study by electron microscopy. J Exp Med 118:327–340.PubMedCrossRefGoogle Scholar
  2. Almeida JD, Waterson AP (1969) The morphology of virus-antibody interaction. Adv Virus Res 15:307–338.PubMedCrossRefGoogle Scholar
  3. Anderson N, Doane FW (1972) Agar diffusion method for negative staining of microbial suspensions in salt solutions. Appl Microbiol 24:495–496.PubMedGoogle Scholar
  4. Anderson N, Doane FW (1973) Specific identification of enteroviruses by immuno-electron microscopy using a serum-in-agar method. Can J Microbiol 19:585–589.PubMedCrossRefGoogle Scholar
  5. Anderson TF, Stanley WM (1941) A study by means of the electron microscope of the reaction between tobacco mosaic virus and its antiserum. J Biol Chem 139:339–344.Google Scholar
  6. Bayer ME, Mannweiler E (1963) Antigen-antibody reactions in influenza virus as seen in the electron microscope. Arch Gesamte Virusforsch 13:541–547.PubMedCrossRefGoogle Scholar
  7. Bayer ME, Blumberg BS, Werner B (1968) Particles associated with Australia antigen in the sera of patients with leukemia, Down’s syndrome and hepatitis. Nature 218:1057–1059.PubMedCrossRefGoogle Scholar
  8. Berthiaume L, Alain R, McLaughlin B, Payment P (1981) Rapid detection of human viruses in faeces by a simple and routine immune electron microscopy technique. J Gen Virol 55:223–227.PubMedCrossRefGoogle Scholar
  9. Best JM, Bantvala JE, Almeida JD, Waterson AP (1967) Morphological characteristics of rubella virus. Lancet 2:237–239.CrossRefGoogle Scholar
  10. Brenner S, Horne RW (1959) A negative staining method for high resolution microscopy of viruses. Biochem Biophys Acta 34:103–110.PubMedCrossRefGoogle Scholar
  11. Derrick KS (1973) Quantitative assay for plant viruses using serological specific electron microscopy. Virology 56:652–653.PubMedCrossRefGoogle Scholar
  12. Doane FW (1974) Identification of viruses by immunoelectron microscopy. In: Kurstak E, Morisset R (eds) Viral Immunodiagnosis. Academic Press, New York, pp 237–255.Google Scholar
  13. Doane FW (1986) Electron microscopy and immunoelectron microscopy. In: Specter S, Lancz GJ (eds) Clinical Virology Manual. Elsevier, New York, pp 71–88.Google Scholar
  14. Doane FW (1987) Immunoelectron microscopy in diagnostic virology. Ultrastruct Pathol 11:681–685.PubMedCrossRefGoogle Scholar
  15. Doane FW (1988a) Immunoelectron microscopy and its role in diagnostic virology. Clin Immunol News 9:159–162.CrossRefGoogle Scholar
  16. Doane FW (1988b) Electron Microscopy. In: Lennette EH, Halonen P, Murphy FA (eds) Laboratory Diagnosis of Infectious Diseases. Principles and Practice. Springer-Verlag, New York, pp 121–131.CrossRefGoogle Scholar
  17. Doane FW, Anderson N (1977) Electron and immunoelectron microscopic procedures for diagnosis of viral infections. In Kurstak E, Kurstak C (eds) Comparative Diagnosis of Viral Diseases, Vol II, part B. Academic Press, New York, pp 505–539.Google Scholar
  18. Doane FW, Anderson N (1987) Electron Microscopy in Diagnostic Virology: A Practical Guide and Atlas. Cambridge University Press, New York.Google Scholar
  19. El-Ghorr AA, Snodgrass DR, Scott FMM (1988) Evaluation of an immunogold electron microscopy technique for detecting bovine coronavirus. J Virol Methods 19:215–224.PubMedCrossRefGoogle Scholar
  20. Faulk WP, Taylor GM (1971) An immunocolloidal method for the electron microscope. Immunochemistry 8:1081–1083.PubMedCrossRefGoogle Scholar
  21. Furui S (1986) Use of protein A in the serum-in-agar diffusion method in immune electron microscopy for detection of virus particles in cell culture. Microbiol Immunol 30:1023–1035PubMedGoogle Scholar
  22. Gerna G, Passarani N, Battaglia M, Percivalle E (1984) Rapid serotyping of human rotavirus strains by solid-phase immune electron microscopy. J Clin Microbiol 19:273–278.PubMedGoogle Scholar
  23. Gerna G, Passarani N, Sarasini A, Battaglia M (1985) Characterization of serotypes of human rotavirus strains by solid-phase immune electron microscopy. J Infect Dis 152:1143–1151.PubMedCrossRefGoogle Scholar
  24. Gerna G, Sarasini A, Coulson BS, Parea M, Torsellini M, Arbustini E, Battaglia M (1988) Comparative sensitivities of solid-phase immune electron microscopy and enzyme-linked immunosorbent assay for serotyping of human rotavirus strains with neutralizing monoclonal antibodies. J Clin Microbiol 26:1383–1387.PubMedGoogle Scholar
  25. Hammond GW, Hazelton PR, Chuang I, Klisko, B. (1981) Improved detection of viruses by electron microscopy after direct ultracentrifuge preparation of specimens. J Clin Microbiol 14:220–221.Google Scholar
  26. Hopley, JFA (1985) Protein A-gold immunoelectron microscopy studies on the simian rotavirus SAH. M.Sc thesis, University of Toronto.Google Scholar
  27. Hopley JFA, Doane FW (1985) Development of a sensitive protein A-gold immunoelectron microscopy method for detecting viral antigens in fluid specimens. J Virol Methods 12:135–147.PubMedCrossRefGoogle Scholar
  28. Horisberger M (1981) Colloidal gold. A cytochemical marker for light and fluorescent microscopy and for transmission and scanning electronmi-croscopy. In: Johari O (ed) Scanning Electron Microscopy. AMF O’Hara, S.E.M., Inc., Chicago, vol II, pp 9–31.Google Scholar
  29. Hummeler K, Anderson TF, Brown RA (1961) Identification of poliovirus particles of different antigenicity by specific agglutination as seen in the electron microscope. Virology 16:84–90.CrossRefGoogle Scholar
  30. Kapikian A, Almeida J, Stott E (1972a) Immune electron microscopy of rhino-viruses. J Virol 10:142–146.PubMedGoogle Scholar
  31. Kapikian AZ, Wyatt RG, Dolin R, Thornhill TS, Kalica AR, Chanock RM (1972b) Visualization by immune electron microscopy of a 27 nm particle associated with acute infectious non-bacterial gastroenteritis. J Virol 10:1075–1081.PubMedGoogle Scholar
  32. Kapikian A, James H, Kelly S, Vaughn A (1973) Detection of coronavirus 692 by immune electron microscopy. Infect Immun 7:111–116.PubMedGoogle Scholar
  33. Kapikian AZ, Dienstag JL, Purcell RH (1976) Immune electron microscopy as a method for the detection, identification, and characterization of agents not cultivable in an in vitro system. In: Rose NR, Friedman H (eds) Manual of Clinical Immunology. Amer Soc Microbiol, Washington D.C. pp 467–480.Google Scholar
  34. Katz D, Straussman Y, Shahar A, Kohn (1980) Solid-phase immune electron microscopy (SPIEM) for rapid viral diagnosis. J Immunol Methods 38:171–174.PubMedCrossRefGoogle Scholar
  35. Katz D, Kohn A (1984) Immunosorbent electron microscopy for detection of viruses. Adv Virus Res 29:169–194PubMedCrossRefGoogle Scholar
  36. Katz D, Straussman Y (1984) Evaluation of immunoadsorbent electron microscopic techniques for detection of Sindbis virus. J Virol Methods 8:243–254.PubMedCrossRefGoogle Scholar
  37. Kelen AE, Hathaway AE, McLeod DA (1971) Rapid detection of Australia/SH antigen and antibody by a simple and sensitive technique of immunoelectron microscopy. Can J Microbiol 17:993–1000.PubMedCrossRefGoogle Scholar
  38. Kjeldsberg E (1985) Specific labelling of human rotaviruses and adenoviruses with gold-IgG complexes. J Virol Methods 12:47–57.PubMedCrossRefGoogle Scholar
  39. Kjeldsberg E (1986) Immunonegative stain techniques for electron microscopic detection of viruses in human faeces. Ultrastr Pathol 10:553–570.CrossRefGoogle Scholar
  40. Kjeldsberg E, Mortensson-Egnund K (1982) Comparison of solid-phase immune electron microscopy, direct electron microscopy and enzyme-linked immunosorbent assay for detection of rotaviruses in faecal samples. J Virol Methods 4:45–53.PubMedCrossRefGoogle Scholar
  41. Kjeldsberg E, Siebke JC (1985) Use of immunosorbent electron microscopy for detection of rota-and hepatitis A virus in sucrose solutions. J Virol Methods 12:161–167.PubMedCrossRefGoogle Scholar
  42. Kleczkowski A (1961) Serological behaviour of tobacco mosaic virus and its protein fragments. Immunol. 4:130–141.Google Scholar
  43. Lafferty KJ, Oertelis SJ (1961) Attachment of antibody to influenza virus. Nature 192:764–765.PubMedCrossRefGoogle Scholar
  44. Lafferty KJ, Oertelis S (1963) The interaction between virus and antibody. III. Examination of virus-antibody complexes with the electron microscope. Virology 21:91–99.CrossRefGoogle Scholar
  45. Lamontagne L, Marsolais G, Marois P, Assaf R (1980) Diagnosis of rotavirus, adenovirus, and herpesvirus infections by immune electron microscopy using a serum-in-agar diffusion method. Can J Microbiol 26:261–264.PubMedCrossRefGoogle Scholar
  46. Lee FK (1977) A study of enterovirus-antibody interaction in immunoelectron microscopy. M.Sc thesis, University of Toronto.Google Scholar
  47. Lin N (1984) Gold IgG complexes improve the detection and identification of viruses in leaf dip preparations. J Virol Methods 8:181–190.PubMedCrossRefGoogle Scholar
  48. Milne RG, Luisoni E (1977) Rapid immune electron microscopy of virus preparations. In: Maramorosch K, Kaprowski H (eds) Methods in Virology. Academic Press, New York, vol 6, pp 265–281.Google Scholar
  49. Morinet F, Ferchal F, Colimon R, Pérol Y (1984) Comparison of six methods for detecting human rotavirus in stools. Eur J Clin Microbiol 3:136–140.PubMedCrossRefGoogle Scholar
  50. Narang HK, Codd AA (1981) Frequency of pre-clumped virus in routine fecal specimens from patients with acute nonbacterial gastroenteritis. J Clin Microbiol 13:982–988.PubMedGoogle Scholar
  51. Nicolaieff A, Obert G, Van Regenmortel MHV (1980) Detection of rotavirus by serological trapping on antibody-coated electron microscope grids. J Clin Microbiol 12:101–104.PubMedGoogle Scholar
  52. Obert G, Gloekler R, Burckard J, Van Regenmortel MHV (1981) Comparison of immunosorbent electron microscopy, enzyme immunoassay and counter-immunoelectrophoresis for detection of human rotavirus in stools. J Virol Methods 3:99–107.PubMedCrossRefGoogle Scholar
  53. Pares RD, Whitecross MI (1982) Gold-labelled antibody decoration (GLAD) in the diagnosis of plant viruses by immune-electron microscopy. J Immunol Methods 51:23–28.PubMedCrossRefGoogle Scholar
  54. Paver W, Ashley C, Caul E, Clarke S (1973) A small virus in human faeces. Lancet 1:237–240.PubMedCrossRefGoogle Scholar
  55. Pegg-Feige (1983) Solid phase immunoelectron microscopy (SPIEM) in virus identification. M.Sc thesis. University of Toronto.Google Scholar
  56. Pegg-Feige K, Doane FW (1983) Effect of specimen support film in solid phase immunoelectron microscopy. J Virol Methods 7:315–319.PubMedCrossRefGoogle Scholar
  57. Pegg-Feige K, Doane FW (1984) Solid-phase immunoelectron microscopy for rapid diagnosis of enteroviruses. Proc of the 42nd Annual Meeting of the Electron Microscopy Society of America, pp 226-227.Google Scholar
  58. Petrovicova A, Juck AS (1977) Serotyping of coxsackieviruses by immune electron microscopy. Acta Virol 21:165–167.PubMedGoogle Scholar
  59. Rubenstein AS, Miller MF (1982) Comparison of an enzyme immunoassay with electron microscopic procedures for detecting rotavirus. J Clin Microbiol 15:938–944.PubMedGoogle Scholar
  60. Stannard LM, Lennon M, Hodgkiss M, Smuts H (1982) An electron microscopic demonstration of immune complexes of hepatitis B e-antigen using colloidal gold as a marker. J Med Virol 9:165–175.PubMedCrossRefGoogle Scholar
  61. Svensson L, Grandien M, Pettersson C-A (1983) Comparison of solid-phase immune electron microscopy by use of protein A with direct electron microscopy and enzyme-linked immunosorbent assay for detection of rotavirus in stool. J Clin Microbiol 18:1244–1249.PubMedGoogle Scholar
  62. van Rij G, Klepper L, Peperkamp E, Schaap GJP (1982) Immune electron microscopy and a cultural test in the diagnosis of adenovirus ocular infection. Brit J Ophthamol 66:317–319.CrossRefGoogle Scholar
  63. von Ardenne M., Friedrich-Freska H, Schramm G (1941) Elektronmikroskopische Untersuchung der pracipitinreaktion von tabakmosaikvirus mit kaninchenantiserum. Arch Gesamte Virusforsch 2:80–86.CrossRefGoogle Scholar
  64. Vreeswijk J, Folkers E, Wagenaar F, Kapsenberg JG (1988) The use of colloidal gold immunoelectron microscopy to diagnose varicella-zoster virus (VZV) infections by rapid discrimination between VZV, HSV-1 and HSV-2. J Virol Methods 22:255–271.PubMedCrossRefGoogle Scholar
  65. Watson DH, Wildy P (1963) Some serological properties of herpes virus particles studied with the electron microscope. Virology 21:100–111.PubMedCrossRefGoogle Scholar
  66. Wood DJ, Bailey AS (1987) Detection of adenovirus types 40 and 41 in stool specimens by immune electron microscopy. J Med Virol 21:191–199.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1991

Authors and Affiliations

  • Frances W. Doane
    • 1
  • Nan Anderson
    • 1
  • Francis Lee
    • 1
  • Kathryn Pegg-Feige
    • 1
  • John Hopley
    • 1
  1. 1.Department of MicrobiologyUniversity of TorontoTorontoCanada

Personalised recommendations