Fig. 3. Validation of BT-AVATar in vitro.

a Titration of BT-AVATar containing dBT-modified PACAP (solid line) showing activation with EC₅₀ of 120 ± 10 nM. An equivalent variant of the sensor containing BT-modified PACAP (dashed line) is not activated by biotin. b Biotin vasosensor activity over repeated cycles of excess SA and biotin (BT) addition. c Schematic showing AVATar activation by biotinylated proteins or cells (red). Inlay shows biotinylated HEK cells stained with fluorescent streptavidin. Scale bar = 50 µm. Experiment was performed twice. d BT-AVATar is robustly activated by biotinylated bovine serum albumin and HEK cells (red) but not by unbiotinylated controls (gray). All error bars denote SD of duplicates.