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An immunofluorescence assay for complement activation by the classical pathway

E Linder, M Rhen, S Meri

    Journal of Immunological Methods
    |January 1, 1981
    PubMed
    Summary
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    This study introduces a new immunofluorescence assay to assess complement pathway function. The assay effectively identifies complement deficiencies by detecting C1q binding to intermediate filaments in placental tissue.

    Area of Science:

    • Immunology
    • Biochemistry

    Background:

    • The classical complement pathway is crucial for immune responses.
    • Assessing complement pathway functional integrity is vital for diagnosing deficiencies.
    • Existing methods can be complex or lack sensitivity.

    Purpose of the Study:

    • To develop and validate a novel immunofluorescence (IFL) assay for evaluating the functional integrity of the classical complement pathway.
    • To utilize cytoskeletal intermediate filaments (IMF) as a substrate for complement activation.
    • To establish a simple method for detecting complement deficiencies.

    Main Methods:

    • An immunofluorescence (IFL) assay was developed using human placental capillary endothelium rich in intermediate filaments (IMF).
    • The assay measures the binding of complement components (C1q, C4, C3) after incubation with human sera.

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  • Inhibitors of C1q binding and C3 convertase formation were used to validate the assay's specificity.

    • Main Results:

    • The IFL assay successfully detected C1q, C4, and C3 binding in normal human sera up to a 1:80 dilution.
    • Known inhibitors blocked complement component binding, confirming assay specificity.
    • Eight out of 100 patient sera showed reduced or negative complement binding, compared to 100% positive control sera from healthy individuals.
    • Further analysis of four deficient sera revealed three with markedly reduced hemolytic activity and normal C3/C4 levels.

    Conclusions:

    • The developed IFL assay is a simple and effective method for assessing classical complement pathway activation.
    • This assay can be used to evaluate complement deficiencies and study complement activation mechanisms.
    • The assay demonstrates potential for clinical diagnostics and research in complement-mediated diseases.