Anti his pe antibody

SpyCatcher-expressing T cells were resuspended in CM containing various concentrations of SpyTag-labeled targeting agent. Cells were incubated for 30–60 min at 37 °C and 5% CO₂ and then washed two times with CM. To stain IgG-SpyTag-armed T cells, goat polyclonal antihuman IgG (Sigma-Aldrich) conjugated with LightningLink APC (Expedeon) was used. To stain myc-or Flag-DARPin-SpyTag-armed T cells, either anti-Myc-Alexa647 (Cell Signaling Technologies) or anti-FLAG-BV421 (BioLegend) was used. Stained cells were analyzed by flow cytometry.
For receptor turnover experiments, T cells were expanded and rested to an average volume of <300 fL and then armed with either SpyTag-labeled IgG or SpyTag-labeled DARPin, as described above. For basal turnover, T cells were maintained in CM and stained for loaded receptor every 24 h for a total of 96 h. For antigen-induced turnover, T cells were combined at a 3:1 effector to target ratio and stained after 24 h in culture.
The expression of 4D5-BBζ and 4D5–28ζ receptors was detected through incubation with soluble Her2-his protein (Sino Biological), followed by staining with anti-His-PE antibody (BioLegend). Her2 expression and EGFR expression were detected using anti-Her2-APC (BioLegend) or anti-EGFR-PE (BioLegend), respectively. Stained cells were analyzed by flow cytometry.