title: Evaluation of complement-dependent cytotoxicity using ATP measurement and C1q/C4b binding.
abstract: One of the major issues for antibody treatment is enhancement of efficacy. Recent studies have highlighted the important role of effector functions in improvement of antibody therapy. Among effector functions, complement-dependent cytotoxicity (CDC), which induces cell lysis by a cascade of activation triggered by the binding of C1q subunits to the Fc regions of antibodies bound to the cell surface, is part of the mechanism of several antibody therapies. CDC can be modulated by either Fc isotype engineering or Fc genetic mutations or Fc glycosylation profile modifications. To evaluate the impact of such modifications on CDC, we describe a luminescence method based on ATP measurement to estimate tumor damaged cells and a flow cytometry method to evaluate the binding of C1q on the Fc region and the binding of C4b on cell surface. The luminescence method coupled with complement protein analysis by flow cytometry encompasses all needed methods to evaluate antibody ability to trigger CDC.
authors.affiliation: Experimental Oncology Department, Centre d'Immunologie Pierre-Fabre, Saint Julien-en-Genevois, France.
authors.forename: Lucile
authors.initials: L
authors.lastname: Broyer
authors.forename: Liliane
authors.initials: L
authors.lastname: Goetsch
authors.forename: Matthieu
authors.initials: M
authors.lastname: Broussas
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D000920:Antibody-Dependent Cell Cytotoxicity; D018951:Antigens, CD20 / Q000276:immunology; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D015922:Complement C1q / Q000378:metabolism*; D015935:Complement C4b / Q000378:metabolism*; D003601:Cytotoxicity Tests, Immunologic / Q000379:methods*; D004354:Drug Screening Assays, Antitumor / Q000379:methods*; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism; D008297:Male; D011485:Protein Binding
chemical_list: D000911:Antibodies, Monoclonal; D018951:Antigens, CD20; D007141:Immunoglobulin Fc Fragments; D015922:Complement C1q; D015935:Complement C4b; D000255:Adenosine Triphosphate