Fig. 7
Binding of geraniin to recombinant DENV-2 E Domain III protein. a SDS-PAGE gel showing expression of DENV-2 E-DIII protein. Lane M indicates the 10 kDa protein ladder, Lane 1- protein induced with IPTG; Lane 2- protein induced with IPTG; Lane 3- uninduced protein. Size of E-DIII protein is 15 kDa. b SDS-PAGE gel showing purification of DENV-2 E-DIII protein. Lane M-10 kDa protein ladder; Lanes; 1-pellet, 2-supernatant, 3-flow-through, 4-wash 1, 5-wash 2, 6-wash 3, 7-eluent. A single band at the expected size indicated that the recombinant protein obtained was pure. c The saturation binding curve obtained when recombinant E-DIII protein was coated onto ELISA plate and was allowed to bind to different concentrations of monoclonal antibody, clone 5j122 (MAB8901) before the signal was detected using an AP conjugated secondary antibody. The Kd value of the mAb was determined from the curve by nonlinear regression using GraphPad Prism 5 software. d The competitive binding curve was obtained through non-linear regression analysis using GraphPad Prism 5 software. The same monoclonal antibody (MAB8901) was used at one constant concentration of 3.33 nM while various concentrations of geraniin was used to test the ability of geraniin to bind to the recombinant E-DIII protein, given that the Kd value of the mAb was known. Data represent mean values ±SEM