Development of a label-free ATP image sensor for analyzing spatiotemporal patterns of ATP release from biological tissues

Abstract We investigated the output characteristics of a label-free, 37.3 µm pitch, 128 × 128 pixel adenosine-5'-triphosphate (ATP) image sensor, in which different thicknesses of ATP-degrading enzyme apyrase were deposited using the poly-ion complex (PIC) method. ATP was detected based on the changes in the concentration of hydrogen ions (H+) generated by enzymatic reactions. Enzyme films of different thicknesses were formed on the surface of the sensor by controlling the volumes of the PIC solution. A reduction in the enzyme film thickness (0.2 µm thickness), improved the sensitivity of the ATP sensor, and a clear image than that obtained with a sensor with a thicker film (2.3 µm thickness) was obtained. Additionally, the detection limit for the 0.2-µm-thick sensor was as low as 10 µM, whereas that of the 2.3-µm thick sensor was 100 µM. To examine ATP release from biological tissues, we placed a mouse hippocampal slice on the sensor with the 0.2-µm-thick film and electrically stimulated the nerve fiber (i.e., Schaffer collateral). The neuronal stimulation evoked significant changes in the output signals of the hippocampal slice but not in the sensor area which was not in contact with tissue. An elevation in the output signal was initiated at the stimulated site with a maximum increment of 5 mV, and the signal traveled along the neuronal fibers. By reducing the film thickness, we successfully developed a high-sensitivity label-free ATPo image sensor with a high spatiotemporal resolution, which allows imaging of endogenous ATPo released from the hippocampus.