Detection of complement activation using monoclonal antibodies against C3d

Area of Science:

• Immunology
• Biochemistry

Background:

• Complement activation involves C3 protein cleavage, generating fragments that bind to tissues.
• Tissue-bound C3 fragments serve as durable biomarkers for inflammation and targets for diagnostics and therapeutics.

Purpose of the Study:

• To generate cross-reactive monoclonal antibodies against human and mouse C3d.
• To evaluate the utility of these antibodies for detecting tissue-bound C3 fragments in disease models.

Main Methods:

• Developed three murine monoclonal antibodies (3d8b, 3d9a, 3d29) targeting C3 activation fragments (iC3b, C3dg, C3d).
• Assessed antibody binding to C3 fragments in solution and on tissues.
• Utilized mouse models of renal and ocular diseases to confirm in vivo antibody accumulation at sites of C3 deposition.
• Employed optical imaging to detect antibody accumulation in retinal lesions in a choroidal neovascularization model.

Main Results:

• The developed antibodies specifically bind to C3 activation fragments but not intact C3 or C3b.
• Systemically injected antibodies accumulated at sites of C3 fragment deposition in renal and ocular tissues.
• Optical imaging successfully detected antibody accumulation within retinal lesions.

Conclusions:

• The generated monoclonal antibodies are effective tools for detecting tissue-bound C3 fragments.
• These antibodies, combined with imaging techniques, offer a sensitive method for monitoring complement activation-associated inflammation.