Commercial horseradish peroxidase degrades myelin encephalitogenic protein during coupling for immunohistochemical studies
Conjugates of myelin encephalitogenic basic protein (EP) and commercial horseradish peroxidase (HRP) have been used for immunohistochemical demonstrations of anti-EP antibody in animals with experimental allergic encephalomyelitis. We performed gel electrophoresis studies on EP-HRP conjugates prepar...
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Published in | The journal of histochemistry and cytochemistry Vol. 25; no. 5; pp. 329 - 336 |
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Main Authors | , |
Format | Journal Article |
Language | English |
Published |
Los Angeles, CA
Histochemical Soc
01.05.1977
SAGE Publications |
Subjects | |
Online Access | Get full text |
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Summary: | Conjugates of myelin encephalitogenic basic protein (EP) and commercial horseradish peroxidase (HRP) have been used for immunohistochemical demonstrations of anti-EP antibody in animals with experimental allergic encephalomyelitis. We performed gel electrophoresis studies on EP-HRP conjugates prepared with glutaraldehyde and on mixtures of EP and HRP incubated without glutaraldehyde. The results show that under conditions of one-and two-step coupling HRP causes rapid loss of the native EP band, apparently due to EP degradation. The EP-HRP mixtures are not encephalitogenic in rabbits, or encephalitogenic activity is lost during processing. The immunohistochemical reactivity of conjugates, however, signals some preservation of antibody-combining sites. The mechanism of the HRP effect on EP is unknown. The possibilities of a contaminating proteinase or direct peroxidatic attack are suggested. Until this action of HRP can be overcome, the effect of coupling procedures on the biological activities of EP will be difficult to assess, and EP-HRP conjugates cannot be expected to reveal sites that may bind encephalitogenic portions of the EP molecule. |
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ISSN: | 0022-1554 1551-5044 |
DOI: | 10.1177/25.5.68067 |