Fluorometric and Flow Cytometric Analysis of the Cyclosporine-Calmodulin Interaction in T Lymphocytes

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Cyclosporine, an intracellular peptide of fungal origin, is a potent immunosuppressive agent. It appears to primarily affect T lymphocyte-mediated immune responses. At the cellular level, cyclosporine appears to interfere with early activation events required for T lymphocyte maturation and proliferation. As calcium-calmodulin-dependent events are shown to play a major role in T lymphocyte activation, the possibility of an interaction between cyclosporine and calmodulin is investigated in this chapter. Analysis was performed at two levels. First, the interaction between these peptides was analyzed in vitro, at the molecular level, using fluorescent derivatives of both calmodulin and cyclosporine. Binding of compounds to these fluorescent derivatives and the conditions of binding are assayed rapidly using fluorometric techniques. Second, the interaction was analyzed in vitro, at the cellular level, using flow cytometric techniques and intact T lymphocytes to analyze the binding conditions of a fluorescent dansylated derivative of cyclosporine. From a number of investigations, it appeared that CsA interferes with early events of T lymphocyte activation to prevent the maturation and proliferation of immunocompetent T lymphocytes. Both calmodulin and the calmodulin inhibitors can competitively inhibit binding of CsA to intact T lymphocytes in a manner similar to native CsA.

Original languageEnglish (US)
Pages (from-to)677-690
Number of pages14
JournalMethods in enzymology
Issue numberC
StatePublished - Jan 1987
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


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