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Methods for Quantitative Detection of Antibody-induced Complement Activation on Red Blood Cells
Published on: January 29, 2014
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Using Bio-Layer Interferometry to Evaluate Anti-PEG Antibody-Mediated Complement Activation.
Mahmoud Mostafa1,2, Nehal E Elsadek1, Sherif E Emam1,3
1Department of Pharmacokinetics and Biopharmaceutics, Institute of Biomedical Sciences, Tokushima University.
Biological & Pharmaceutical Bulletin
|January 4, 2022
Summary
Bio-layer interferometry (BLI) offers a simpler method to detect anti-polyethylene glycol (PEG) IgM-mediated complement activation, crucial for understanding the accelerated blood clearance (ABC) phenomenon in nanomedicine. This technique successfully identified complement activation by anti-PEG IgM antibodies.
Area of Science:
- Biotechnology
- Immunology
- Nanomedicine
Background:
- Complement activation significantly influences the clearance of PEGylated nanomedicines.
- The accelerated blood clearance (ABC) phenomenon, involving anti-PEG antibodies, is linked to complement system activity.
- Accurate detection of anti-PEG IgM-mediated complement activation is vital for nanomedicine safety and efficacy.
Purpose of the Study:
- To develop and validate a Bio-layer interferometry (BLI) system as an alternative method.
- To directly evaluate anti-polyethylene glycol (PEG) immunoglobulin M (IgM)-mediated complement activation.
- To assess the role of complement in the accelerated blood clearance (ABC) phenomenon.
Main Methods:
- Immobilization of mPEG2000-DSPE antigen onto BLI biosensor chips.
- Incubation with different sources of anti-PEG IgM antibodies (monoclonal and polyclonal).
- Measurement of signal changes indicative of antibody binding and complement activation using BLI.
- Confirmation of results using conventional enzyme-linked immunosorbent assay (ELISA).
Main Results:
- BLI system detected increased signals corresponding to anti-PEG IgM binding to the immobilized antigen.
- Further signal enhancement was observed upon incubation with active mouse serum, indicating complement activation.
- No significant signal increase occurred with heat-inactivated serum, confirming complement-dependent activation.
- BLI findings were consistent with conventional ELISA results for complement activation.
Conclusions:
- Bio-layer interferometry (BLI) is a viable and simple alternative for detecting anti-PEG IgM-mediated complement activation.
- The developed BLI system effectively measures complement activation relevant to the ABC phenomenon.
- This method provides a direct approach for evaluating immune responses to PEGylated nanomedicines.
