Extended Data Figure 2. Characterization of aY-modified G-GECO1 (Ca²⁺ sensor).

(a) Fluorescence excitation (dash line) and emission (solid line) profiles for G-GECO1 after addition of 1 mM EGTA (cyan) or 100 μM Ca²⁺ (dark cyan), and aY-G-GECO1 after addition of 1 mM EGTA (magenta) or 100 μM Ca²⁺ (red). (b) Representative images of HeLa cells expressing aY-G-GECO1 in response to sequential addition of 5 μM histamine, 1 mM CaCl₂ with 10 μM ionomycin, and 2 mM EGTA with 5 μM ionomycin. Scale bar: 30 μm. (c) Quantitative traces for randomly selected five cells in panel b. Intensities are normalized to the values at t = 0 s. The time points for addition of chemicals are shown as arrows. Ca²⁺ oscillations in response to histamine were observed as expected. These experiments were repeated three times with similar results using independent biological samples.