25ng of hACE2-Fc fusion protein expressed in HEK293 cells were adhered to high-capacity binding, 96 well-plates (Corning) overnight in PBS. Plates were blocked with 5% BSA in PBS containing Tween-20 (PBS-T) for 1 h at room temperature (RT). Blocking solution was discarded and 5-fold dilutions of 6xHis-tagged RBDs in PBS were added to wells and incubated for 1 h at RT. Plates were then washed three times with PBS-T. Anti-polyhistidine IgG-Biotin (Abcam) in PBS-T was added to each and incubated for 1 h at RT. Plates were then washed three times with PBS-T. Streptavidin-HRP (horseradish peroxidase) (Abcam) in PBS-T was added to each and incubated for 1 h at RT. Plates were then washed three times with PBS-T Plates were developed using 1-Step Ultra TMB (3,3′,5,5′-tetramethylbenzidine) substrate (ThermoFisher), stopped with sulfuric acid and immediately read using a plate reader at 450nm. Data were plotted using Prism 9 (GraphPad Software) and affinities determined by applying a nonlinear regression model.
Anti polyhistidine igg biotin
Manufactured by Abcam
Anti-polyhistidine IgG-Biotin is a biotinylated antibody that binds to polyhistidine tags. It can be used in various applications, such as affinity purification or detection of proteins with polyhistidine tags.
Automatically generated - may contain errors
2 protocols using anti polyhistidine igg biotin
1
SARS-CoV-2 RBD Binding Assay
2
Quantifying SARS-CoV-2 RBD-hACE2 Interactions
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25ng of hACE2-Fc fusion protein expressed in HEK293 cells were adhered to high-capacity binding, 96 well-plates (Corning) overnight in PBS. Plates were blocked with 5% BSA in PBS containing Tween-20 (PBS-T) for 1hr at room temperature (RT). Blocking solution was discarded and 5-fold dilutions of 6xHis-tagged RBDs in PBS were added to wells and incubated for 1hr at RT. Plates were then washed three times with PBS-T. Anti-polyhistidine IgG-Biotin (Abcam) in PBS-T was added to each and incubated for 1hr at RT. Plates were then washed three times with PBS-T. Streptavidin-HRP (Abcam) in PBS-T was added to each and incubated for 1hr at RT. Plates were then washed three times with PBS-T Plates were developed using 1-Step Ultra TMB substrate (ThermoFisher), stopped with sulfuric acid and immediately read using a plate reader at 450nm. Data were plotted using Prism 9 (GraphPad Software) and affinities determined by applying a nonlinear regression model.
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