Abstract
The peroxidase-antiperoxidase (PAP) method was pioneered by Sternberger in 1979 (1). The method uses an immunological sandwich amplification and the enzyme peroxidase to effect a signal. The unique feature of this procedure is the enzyme-antibody solution, the PAP immune complex. The horseradish peroxidase enzyme, itself an immunogenic protein, is used to inoculate a given species, and a polyclonal immune response is generated against the enzyme. This antiserum is harvested and placed in solution with the enzyme so that immune complexes form that remain soluble. These complexes form with a molar ratio of two molecules of IgG to three molecules of enzyme. Furthermore, not only does this complex remain soluble, but the enzymatic activity of the peroxidase is not affected by the attached immunoglobulins. The antiperoxidase antibodies are from the same species as that which has produced a primary antibody raised against a tissue antigen. These two antibodies, one directed against the tissue antigen and the other directed against the peroxidase enzyme, can be linked by another antibody raised in an alternate species against immunoglobulin from the first species (see Fig. 1).
The opmions or assertions herem represent the personal views of the author, and are not to be construed as offictal or as representmg the views of the Department of the Army or the Department of Defense
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Sternberger, L. (1979) Immunocytochemistry, 2nd ed. Wiley, New York.
Cerio, R. and MacDonald, D (1988) Routine diagnostic immunohistochemical labeling of extracellular antigens in formol saline solution—fixed, paraffinembedded cutaneous tissue J. Am. Acad. Dermatol. 19, 747–753.
Clements, J and Beitz, A. (1985) The effects of different pretreatment conditions and fixation regimes on serotonin immunoreactivity: a quantitative light microscopic study. J. Histochem. Cytochem. 33, 778–784
Laitinen, L., Laitinen, A, Panula, P., Partanen, M., Tervo, K., and Tervo, T. (1983) Immunohistochemical demonstration of substance P in the lower respiratory tract of the rabbit and not of man Thorax 38, 531–536.
Hsu, S. and Soban, E (1982) Color modification of diaminobenzidine (DAB) precipitation by metallic ions and its application for double immunohistochemistry. J. Histochem. Cytochem 30, 1079
Ordronneau, P., Lindstrom, P., and Petrusz. P. (1981) Four unlabeled antibody bridge techniques: A comparison. J. Histochem. Cytochem. 29, 1397–1404.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1994 Humana Press Inc.
About this protocol
Cite this protocol
Bratthauer, G.L. (1994). The Peroxidase-Antiperoxidase (PAP) Method. In: Javois, L.C. (eds) Immunocytochemical Methods and Protocols. Methods in Molecular Bilogy, vol 34. Humana Press. https://doi.org/10.1385/0-89603285-X:165
Download citation
DOI: https://doi.org/10.1385/0-89603285-X:165
Publisher Name: Humana Press
Print ISBN: 978-0-89603-285-9
Online ISBN: 978-1-59259-521-1
eBook Packages: Springer Protocols