Non-cross-reactive anti IgG antibodies

#This application is a 371 of Application No. PCT/EP2010/065617, filed Oct. 18, 2010, which claims priority to European Application number 09013144.2 filed Oct. 19, 2009.

#Herein are reported antibodies specifically binding to the constant region of an IgG-Fab-fragment of human and chimpanzee antibodies of the IgG class and use thereof in immunoassays.

#Since the development of the first monoclonal antibodies by Koehler and Milstein in 1974, a lot of efforts have been dedicated to the development of antibodies which are appropriate for therapy in humans. The first monoclonal antibodies which became available had been developed in mice and rats. In the past ten years an ever growing number of human monoclonal antibodies or humanized monoclonal antibodies have reached the market. Well-known examples include for example Herceptin® and MabThera® from F. Hoffmann-La Roche AG, Basel.

#FIGS. 1a)-1c) Fully generic Assay for quantification of human antibodies (human IgG) in an experimental animal: FIG. 1a) assay format; FIG. 1b) capture and detection reagent: antibody M-R10Z8E9; FIG. 1c) capture and detection reagent antibody M-1.7.10; therapeutic antibodies: empty triangles: anti-IL13Rα1 antibody, empty squares: anti-Abeta antibody, solid squares: anti-IL1R antibody, solid triangles anti-IL6R antibody.

#FIG. 2 Results obtained with an assay employing the antibodies as reported herein; antibodies used from left to right: M-R10Z8E9, M-1.3.2, M-1.5.8, M-1.7.10.

#FIGS. 3a)-3c) Exemplary surface plasmon surface resonance diagrams of antibodies: FIG. 3a) M-1.3.2, FIG. 3b) M-1.5.8, and FIG. 3c) M-1.7.10 as reported herein.

#Preparation of the F(Ab′)2 Fragment of Human IgG (Immunogen)

#The full length human antibody of the class G (human IgG) in 100 mM sodium citrate buffer, pH 3.7 was incubated with pepsin (1 μg pepsin per mg IgG). The fragmentation was analyzed by analytical gel filtration and stopped after 90 minutes by adjusting the pH value to 6.5 by the addition of potassium phosphate. After dialysis of the mixture against 10 mM sodium citrate buffer with 10 mM sodium chloride, pH 5.5, the solution was applied to an SP-Sepharose® chromatography column and the isolated fractions eluted in a salt gradient were analyzed individually by analytical gel filtration. The pool containing the antibody F(ab′)2 fragments were applied to an affinity matrix with immobilized polyclonal antibodies against human Fcγ to eliminate trace amounts of Fcγ fragments. The flow through was pooled, concentrated to about 16 mg/ml and finally applied to a gel filtration column (Superdex® 200).

#Generation of Monoclonal Anti-Human IgG Antibodies