Related Experiment Videos
A C3 convertase assay for nephritic factor functional activity
E Jelezarova1, M Schlumberger, S Sadallah
1Institute of Biochemistry, Swiss Federal Institute of Technology, ETH-Zentrum, CH-8092, Zurich, Switzerland.
Journal of Immunological Methods
|April 9, 2001
Summary
C3 nephritic factor (C3NeF) stabilizes the C3 convertase, driving alternative pathway activation and C3 depletion. A new solid-phase assay quantifies this stabilizing activity for C3NeF IgG.
Area of Science:
- Immunology
- Complement System Biology
Background:
- C3 nephritic factor (C3NeF) is an autoantibody targeting the C3 convertase.
- C3NeF stabilizes the C3 convertase, leading to continuous alternative pathway activation and C3 depletion.
- NeF is associated with membranoproliferative glomerulonephritis and partial lipodystrophy.
Purpose of the Study:
- To develop a reproducible solid-phase assay for quantifying the functional activity of purified C3NeF IgG.
- To characterize the stabilizing activity of C3NeF IgG on C3 convertase decay.
Main Methods:
- Generated C3 convertase on immobilized C3b or C3b(2)-IgG complexes.
- Assessed C3 convertase decay in the presence of normal IgG or NeF IgG, with or without Factor H.
- Measured residual convertase activity using radiolabeled C3 and a capture system.
Main Results:
- Developed a solid-phase assay to measure C3NeF IgG functional activity.
- Demonstrated dose-dependent stabilizing activity of NeF IgG on C3 convertase decay, even with Factor H present.
- Observed similar NeF IgG stabilizing activity using both C3b and C3b(2)-IgG complexes.
Conclusions:
- The developed solid-phase assay provides reproducible data on C3NeF IgG functional activity.
- C3NeF IgG stabilizes the C3 convertase, confirming its role in alternative pathway dysregulation.
- The assay is applicable to purified C3NeF IgG, including that isolated from urine.