Electrochemiluminescence imaging of respiratory activity of cellular spheroids using sequential potential steps

The respiratory activity of cultured cells can be electrochemically monitored using scanning electrochemical microscopy (SECM) with high spatial resolution. However, in SECM, the electrode takes a long time to scan, limiting simultaneous measurements with large biological samples such as cell spheroids. Therefore, for rapid electrochemical imaging, a novel strategy is needed. Herein, we report electrochemiluminescence (ECL) imaging of spheroid respiratory activity for the first time using sequential potential steps. L-012, a luminol analog, was used as an ECL luminophore, and H₂O₂, a sensitizer for ECL of L-012, was generated by the electrochemical reduction of dissolved O₂. The ECL imaging visualized spheroid respiratory activity-evidenced by ECL suppression-corresponding to O₂ distribution around the spheroids. This method enabled the time-lapse imaging of respiratory activity in multiple spheroids with good spatial resolution comparable to that of SECM. Our work provides a promising high-throughput imaging strategy for elucidating spheroid cellular dynamics.