preTCR complexes can traffic to the cell surface in HEK cells. (A) Schematic describing the internalization assay, where antibody binding to cell surface receptors can be quantified through the time-dependent increase in cell fluorescence when labeled antibodies become internalized. PM, plasma membrane; ER, endoplasmic reticulum. (B) Raw flow cytometry plots of one repeat of the internalization assay applied to αβTCR and preTCR-expressing HEK cells. The time-dependent increase in fluorescence intensity is plotted against receptor expression (quantified by TCRβ-GFP). (C) Quantification of internalization assay with cellular fluorescence plotted as a function of both time and receptor expression. Dataset is representative of two biological replicates.