The 96- well maxisorp plates (Thermo Scientific, USA) were coated overnight with 5μg/ml of 14c10 hG1. The antibodies, E1 mG2a and E1 mG2a/hG3 were used at a starting concentration of 5μg/ml and serially diluted half log in blocking buffer (4% MPBS). The binding of the antibodies was detected with anti-mouse-IgG-Fc-HRP (1:5000) (Thermo Scientific, USA) and the reaction was developed with TMB substrate and stopped with 1M sulphuric acid. All incubation steps lasted for one hour at room temperature. Plate washing steps were the same as polyclonal phage ELISA.
Anti mouse igg fc hrp
Manufactured by Thermo Fisher Scientific
Sourced in United States
Anti-mouse-IgG-Fc-HRP is a secondary antibody conjugated with horseradish peroxidase (HRP). It is used to detect the presence of mouse immunoglobulin G (IgG) in a sample during immunoassay procedures.
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2 protocols using anti mouse igg fc hrp
1
Antibody Binding Assay Using ELISA
2
CRM 197 Protein Analysis by SDS-PAGE
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The CRM 197 protein (10 μg) was treated with 0.1 μg trypsin at 37ºC for 5 min and analyzed by 10% SDS-PAGE under reduced and non-reduced conditions. The protein bands were transferred to a nitrocellulose membrane (GE Healthcare, France). Then the membrane was incubated with 3F9 mAb (1 μg/ml) or mouse anti-CRM 197 polyclonal antibody (1:2,000 v/v) diluted in blocking solution (5% skim milk in 0.05% TBST), followed by anti-human IgG Fc-HRP conjugate (1:6,000 v/v, Thermo Fisher Scientific) or anti-mouse IgG Fc-HRP (1:5,000 v/v, Thermo Fisher Scientific), respectively. Finally, the bands were visualized using a chemiluminescent substrate (WEST-ZOL plus, iNtRON BioTechnology, Korea).
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