Accurately measuring respiratory activity of single living cells by scanning electrochemical microscopy

Scanning electrochemical microscopy (SECM) is a powerful tool to examine the respiratory activity of living cells. However, in SECM measurements of cell respiratory activity, the signal recorded usually also includes the signal corresponding to the cell topography. Therefore, measurements of cell respiratory activity using conventional SECM techniques are not accurate. In the present work, we develop a method for accurate measurement of the respiratory activity of single living cells using SECM. First, cells are immobilized on a glass substrate modified with collagen. Then, a Pt ultramicroelectrode tip of SECM held at −0.50 V is scanned along the central line across a living cell and a SECM scan curve, i.e., the relationship of the tip current versus the displacement (the first scan curve) is recorded with a negative peak. The peak current ip on this first scan curve is composed of ip1, which corresponds to the cell respiratory activity and ip2, which corresponds to the cell topography. In order to isolate the ip2 component, the cell is killed by exposing it to 1.0 × 10−3 mol/L KCN for 10 min. The tip is then scanned again with the same trace over the dead cell, and a second SECM scan curve is recorded. Noting that the topography of the dead cell is the same as that of the living cell, this second scan curve with a negative peak corresponds now only to the cell topography. Thus, ip2 is obtained from the second SECM scan curve. Finally, ip1 corresponding to the respiratory activity of the living cell can be accurately calculated using ip1 = ip − ip2. This method can be used to monitor real-time change in the respiratory activity of single cells after exposing them to KBr, NaN3 and KCN.