2.5. Depth of puncture

A confocal laser scanning microscope (A1RMP, Nikon, Tokyo, Japan) was used to obtain images of the MNs puncturing the skin. The excitation and emission wavelengths of the FR were 510 nm and 600 nm, respectively. A laser with a wavelength of 530 nm was used to excite FR. The skin of an isolated porcine ear was used as a model skin [35]. The skin was scanned at a frequency of 20 µm per time. The 3D reconstructed images were obtained using XYZ stacking.