Horseradish Peroxidase Labeling of Antibody Using Periodate Oxidation

  • G. Brian Wisdom
Part of the Springer Protocols Handbooks book series (SPH)


The most commonly used method (1) for labeling IgG antibody molecules with horseradish peroxidase exploits the glycoprotein nature of the enzyme. The saccharide residues are oxidized with sodium periodate to produce aldehyde groups that can react with the amino groups of the IgG molecule, and the Schiff bases formed are then reduced to give a stable conjugate of high molecular weight (0.5–1 × 106). The peroxidase has few free amino groups, so self-coupling is not a significant problem.


Horseradish Peroxidase Schiff Base Sodium Acetate Buffer Sodium Borohydride Sodium Periodate 
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  1. 1.
    Wilson, M. B. and Nakane, P. P. (1978) Recent developments in the periodate method of conjugating horseradish peroxidase (HRPO) to antibodies, in Immunofluorescence and Related Staining Techniques (Knapp, W., Holubar, K., and Wick, G., eds.), Elsevier, North Holland Biomedical, Amsterdam, pp. 215–224.Google Scholar
  2. 2.
    Maseyeff, R., Maiolini, R., Ferrua, B., and Ragimbeau-Gilli, J. (1976) Quantitation of alpha fetoprotein by enzyme immunoassay, in Protides of the Biological Fluids, Proc. 24th Colloquium (Peeters, H., ed.), Pergamon, Oxford, pp. 605–612.Google Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 1996

Authors and Affiliations

  • G. Brian Wisdom
    • 1
  1. 1.Division of Biochemistry, School of Biology and BiochemistryThe Queen’s University, Medical Biology CentreBelfastUK

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