Figure 5.

Flow cytometry-based experiments identify ATP1B2 as a target for ACSA-2. HEK293T cells (which do not bind ACSA-2 under normal conditions) were transfected with plasmids encoding for proteins identified by our bioinformatic screen (see Table 3). These plasmids also expressed soluble GFP as a marker for successful plasmid transfection. Cells were then stained with an ACSA-2-APC conjugate and analyzed by flow cytometry. From the list of proteins identified in Table 3, only cells expressing ATP1B2 showed strong co-labeling for ACSA-2 and GFP (28.2% of cells). A representative experiment is presented in the figure. This experiment was repeated twice using independent samples (effectively 1 technical replicate per sample) on separate days with the same results. At least 60,000 cells were analyzed per sample. Lines in each plot delineate gates; numbers represent the proportion of cells in each particular gate.