Estimation of immunoglobulin protease activity by quantitative rocket immunoelectrophoresis

Maureen O. Lassiter, Janice C. Kindle, Laura C. Hobbs, Richard L. Gregory

Research output: Contribution to journalArticle

3 Scopus citations

Abstract

Previous methods for estimating immunoglobulin protease activity have involved the use of enzyme-linked immunosorbent assays (ELISA) or sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by autoradiography or Western blotting techniques. An alternative method has been developed to estimate proteolytic activity on human IgA1 and IgG using quantitative rocket immunoelectrophoresis. The method uses agarose containing anti-human IgA or anti-human IgG heavy chain-specific reagent to which protease-digested human immunoglobulin samples are applied to wells and electrophoresed overnight. Because proteolytic activity of immunoglobulins results in many smaller fragments, the optimal antigen-antibody ratio for precipitation changes and migration in an electric field results in a larger rocket. Consequently, the area of the rocket will be larger in a protease-treated immunoglobulin sample than a saline-treated immunoglobulin control. These increased rocket areas are correlated with our ELISA protease results (r≥0.90), as well as with our immunoblot results. The method is sensitive to increasing exposure to proteolysis, as well as to increasing amounts of protease. This technique can be used to quickly estimate the ability of a sample to cleave immunoglobulins.

Original languageEnglish (US)
Pages (from-to)63-69
Number of pages7
JournalJournal of Immunological Methods
Volume123
Issue number1
DOIs
StatePublished - Sep 29 1989
Externally publishedYes

Keywords

  • Immunoglobulin
  • Protease
  • Rocket immunoelectrophoresis

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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