Monoclonal C5 antibody clone 4G2 recognizes human, rabbit and rat complement C5.
The complement system plays important roles in both innate and adaptive immune response and can produce an inflammatory and protective reaction to challenges from pathogens. Complement C5 is a central molecule in all three pathways and after cleavage by its convertases, it initiates the terminal pathway in order to generate the cytolytic MAC.
C5 is mainly synthesised in the liver as a single polypeptide chain and is present in serum in a concentration of 50-80 µm/ml. Besides, local synthesis of C5 is also supported by other cell types including monocytes/macrophages, neutrophils, fibroblasts, and astrocytes. Before secretion the molecule is glycosylated and secreted into plasma as a 190 kDa glycoprotein consisting of a disulphide linked alpha-chain (111 kDa) and beta-chain (75 kDa).
The complement has become an interesting therapeutic target. Especially after the success of Eculizumab, a monoclonal antibody against C5, in the treatment for aHUS and PNH and the application in clinical trials for many other diseases. Monoclonal antibody 4G2 acts as an equivalent of Eculizumab and recognizes human and rabbit C5, however the affinity for rat C5 is even stronger. This cross reactivity makes it a powerful tool in translational animal studies. Besides C5 the antibody also recognizes C5b6, but not C6. The antibody recognizes intact alpha chain and binds to an epitope distinct from Eculizumab. Using haemolytic assays the antibody shows efficient inhibition of AP as well as CP, in all mentioned models.
Clone 4G2 shows strong binding to C5 with a relatively slow off rate in rat. Suggesting to be functional in in vivo studies. Indeed, 4G2 was applied in a rat prophylactic Myasthenia gravis model. In this study the animals were protected from disease and for destruction of the muscle endplates.
Antibody 4G2 has been tested in western blotting, ELISA, and functional studies (Ref. 1)
1) Zelek_W et al; Development and characterization of novel anti-C5 monoclonal antibodies capable of inhibiting complement in multiple species; Immunology, 157, 283–295
Functional studies, Immuno assays, Western blot
W: A non-reduced sample treatment and SDS-PAGE was used. The band size is 110 kDa (Ref.1)
FS: Antibody 4G2 functions as an inhibiting antibody (Ref.1). The 50% complement inhibitory dose is dependent on the type of assay (e.g. haemolysis assay or pathway ELISA) and should be determined per institute in combination with a proper reference control.
Dilutions to be used depend on detection system applied. It is recommended that users test the reagent and determine their own optimal dilutions. The typical starting working dilution is 1:50.