A comparison of the immunogenicity of the native and denatured forms of a protein.
The effect of heat denaturation on the physicochemical and immunological properties of a model protein, ovalbumin, and its formaldehyde/lysine-treated form was investigated. Polyacrylamide gel electrophoresis and gel filtration showed that heat denaturation converted ovalbumin to high Mr polymers, whereas formaldehyde/lysine-treated ovalbumin remained monomeric with only a small proportion forming oligomers. NMR analysis demonstrated that non-denatured structures could easily be differentiated from the denatured structures. Intraperitoneal immunization of rabbits and mice showed that both native and denatured forms of ovalbumin induced an immune response, but denatured forms of ovalbumin were found to be less immunogenic and to have a lower epitope density than native ovalbumin. Analysis of the antisera in crossed immunoelectrophoresis showed that they were specific for either native or denatured forms of ovalbumin. These findings were further investigated by ELISA and immunoaffinity chromatography, and the high specificity and low cross-reactivity was confirmed. We conclude that the immunogenic epitopes on denatured ovalbumin are different from those on ovalbumin, and that these epitopes reflect a continuum of denatured conformations.
Koch
C
C
Centre for Biomolecular Recognition, Department of Immunology, Statens Seruminstitut, Copenhagen, Denmark.
Jensen
S S
SS
Oster
A
A
Houen
G
G
D000818:Animals; D000917:Antibody Formation; D000941:Antigens / Q000737:chemistry; D002846:Chromatography, Affinity; D003429:Cross Reactions; D003433:Crosses, Genetic; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis; D006358:Hot Temperature; D007123:Immunoelectrophoresis, Two-Dimensional; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008805:Mice, Inbred A / Q000276:immunology; D008807:Mice, Inbred BALB C / Q000276:immunology; D008815:Mice, Inbred Strains / Q000276:immunology; D010047:Ovalbumin / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification; D011489:Protein Denaturation / Q000276:immunology*; D011817:Rabbits / Q000276:immunology; D014508:Urea
D000941:Antigens; D000939:Epitopes; D014508:Urea; D010047:Ovalbumin