Application Notes

Multiscale Simultaneous ER and Cytosolic Ca²⁺ Imaging in Plants

July 2026

Calcium ions (Ca²⁺) play a central role in cellular signaling, with environmental and developmental stimuli triggering specific increases in cytosolic and endoplasmic reticulum (ER) Ca²⁺ levels. To define the role of the ER in regulating cytosolic Ca²⁺ dynamics, it is essential to analyse Ca²⁺ fluctuations in both compartments in vivo. This is now feasible using genetically encoded fluorescent Ca²⁺ indicators (GECIs) such as ER‑GCaMP6‑210 and R‑GECO1, which form an ideal pair for simultaneous ER and cytosolic imaging. Here, we describe an Arabidopsis thaliana transgenic line co‑expressing these indicators, enabling visualization of Ca²⁺ responses to stimuli including leaf wounding and extracellular ATP. Our approach allows Ca²⁺ imaging across spatial scales: the Nikon SMZ18 stereomicroscope supports whole‑organ observations in adult leaves, while the AX R laser‑scanning confocal microscope with silicon objectives enables high‑resolution, dual‑compartment analyses at the single‑cell level in roots. Together, these experiments demonstrate that this dual‑sensor line provides a robust platform for investigating ER‑related Ca²⁺ signaling across developmental stages.

Keywords: Plant Biology, Calcium imaging, Arabidopsis thaliana, confocal microscope, stereo microscope