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Development of an assay of neutralizing antibodies to the complement inhibitor CDX‐1135
Author(s) -
Gergel Lauren E.,
Pilsmaker Catherine D.,
Forsberg Eric M.,
Vitale Laura,
Keler Tibor,
Marsh Henry C.,
Thomas Lawrence J.
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.lb591
Subject(s) - complement system , antibody , chemistry , neutralizing antibody , complement receptor 1 , alternative complement pathway , complement receptor , classical complement pathway , microbiology and biotechnology , monoclonal antibody , microtiter plate , recombinant dna , ic3b , immunology , biology , biochemistry , gene
CDX‐1135 (sCR1) is a soluble recombinant glycoprotein derived from the human complement regulatory protein Complement Receptor Type 1 (CR1, CD35, C3b/C4b receptor). CDX‐1135 inhibits the classical, alternative and lectin pathways of the complement cascade, and is being evaluated for use in certain rare renal diseases involving dysregulated complement. A theoretical possibility exists that humans receiving CDX‐1135 may make neutralizing antibodies to this molecule. To monitor for this, we developed a CDX‐1135 neutralizing antibody assay. Briefly, a microtiter plate is coated with S. enteritidis LPS and test samples and controls, heat‐inactivated as needed, are serially diluted in assay buffer that selectively allows alternative complement activation. CDX‐1135 and normal human serum are added to the plate followed by mouse anti‐human C5b‐9 mAb to detect the terminal complement complex (TCC). Peroxidase conjugated F(ab’)2 anti‐mouse IgG in then added. The plate is developed with TMB substrate and endpoint titers are calculated from the baseline patient serum of the same dilution. In conclusion, this assay assesses the neutralizing activity of anti‐CDX‐1135 antibodies on the capacity of CDX‐1135 to inhibit the alternative complement pathway.