Binding of chicken anti-Adalimumab idiotype to Adalimumab vs human IgG1

Figure 1, Binding of anti-Adalimumab is only to the idiotype and not the whole IgG molecule. Anti Adalimumab idiotype was tested for binding to Adalimumab and human IgG1 by indirect ELISA. Adalimumab or human IgG1 was coated onto ELISA plates at equivalent concentrations and blocked using 2% BSA in PBS. Chicken anti-Adalimumab idiotype was diluted to 125ng/ml and applied in triplicate to the coated wells and incubated for 1 hour at room temperature. After washing the wells, peroxidase conjugated detection antibody (Rabbit anti chicken IgY) was applied to the wells at a concentration of 5ng/ml and incubated for 1 hour at room temperature. After a final wash, the wells were developed with TMB and absorbance at 450nm was measured. These results show that the anti-Adalimumab idiotype antibody is strongly specific to the Adalimumab binding site (idiotype), and has negligible binding to the conserved regions of the IgG1 protein.

Comparing chicken anti-Adalimumab idiotype to monoclonal  anti-Adalimumab idiotype in Adalimumab sandwich ELISA.

Figure 2. Comparison of a chicken anti-Adalimumab idiotype and monoclonal anti-Adalimumab idiotype for use in Adalimumab capture ELISA. Each antibody (chicken anti-Adalimumab idiotype vs. monoclonal antibody) was coated onto polystyrene ELISA plates at equivalent concentrations for comparison in a Adalimumab capture ELISA. Adalimumab was spiked into human serum at concentrations from 25 ng/ml down to 1.56ng/ml and applied to each ELISA plate in duplicate and incubated for 1 hour at room temperature. After washing, peroxidase conjugated detection antibody (anti-human IgG1) was applied to the wells at a concentration of 650pg/ml and incubated for 1 hour at room temperature. After a final wash, the wells were developed with TMB and absorbance at 450nm was measured. These results show that the chicken anti-Adalimumab idiotype antibody performs better as a capture antibody than the competing monoclonal antibody and provides higher resolution in the tested range.