title: Detection of complement activation using monoclonal antibodies against C3d.
abstract: During complement activation the C3 protein is cleaved, and C3 activation fragments are covalently fixed to tissues. Tissue-bound C3 fragments are a durable biomarker of tissue inflammation, and these fragments have been exploited as addressable binding ligands for targeted therapeutics and diagnostic agents. We have generated cross-reactive murine monoclonal antibodies against human and mouse C3d, the final C3 degradation fragment generated during complement activation. We developed 3 monoclonal antibodies (3d8b, 3d9a, and 3d29) that preferentially bind to the iC3b, C3dg, and C3d fragments in solution, but do not bind to intact C3 or C3b. The same 3 clones also bind to tissue-bound C3 activation fragments when injected systemically. Using mouse models of renal and ocular disease, we confirmed that, following systemic injection, the antibodies accumulated at sites of C3 fragment deposition within the glomerulus, the renal tubulointerstitium, and the posterior pole of the eye. To detect antibodies bound within the eye, we used optical imaging and observed accumulation of the antibodies within retinal lesions in a model of choroidal neovascularization (CNV). Our results demonstrate that imaging methods that use these antibodies may provide a sensitive means of detecting and monitoring complement activation-associated tissue inflammation.
authors.affiliation: Department of Medicine, University of Colorado School of Medicine, Aurora, Colorado 80045, USA. Joshua.Thurman@ucdenver.edu
authors.forename: Joshua M
authors.initials: JM
authors.lastname: Thurman
authors.forename: Liudmila
authors.initials: L
authors.lastname: Kulik
authors.forename: Heather
authors.initials: H
authors.lastname: Orth
authors.forename: Maria
authors.initials: M
authors.lastname: Wong
authors.forename: Brandon
authors.initials: B
authors.lastname: Renner
authors.forename: Siranush A
authors.initials: SA
authors.lastname: Sargsyan
authors.forename: Lynne M
authors.initials: LM
authors.lastname: Mitchell
authors.forename: Dennis E
authors.initials: DE
authors.lastname: Hourcade
authors.forename: Jonathan P
authors.initials: JP
authors.lastname: Hannan
authors.forename: James M
authors.initials: JM
authors.lastname: Kovacs
authors.forename: Beth
authors.initials: B
authors.lastname: Coughlin
authors.forename: Alex S
authors.initials: AS
authors.lastname: Woodell
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Pickering
authors.forename: Bärbel
authors.initials: B
authors.lastname: Rohrer
authors.forename: V Michael
authors.initials: VM
authors.lastname: Holers
mesh_terms: D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000276:immunology*; D015415:Biomarkers / Q000378:metabolism; D020256:Choroidal Neovascularization / Q000378:metabolism; D003167:Complement Activation; D050577:Complement C3-C5 Convertases / Q000276:immunology; D015933:Complement C3d / Q000276:immunology* / Q000502:physiology; D000939:Epitopes / Q000276:immunology; D006801:Humans; D007249:Inflammation; D051379:Mice; D008810:Mice, Inbred C57BL; D011485:Protein Binding; D011994:Recombinant Proteins / Q000276:immunology; D013154:Spleen / Q000166:cytology; D020349:Surface Plasmon Resonance
chemical_list: D058846:Antibodies, Monoclonal, Murine-Derived; D015415:Biomarkers; D000939:Epitopes; D011994:Recombinant Proteins; D015933:Complement C3d; D050577:Complement C3-C5 Convertases