Principle of Spinning Disk
The illumination source used is usually a laser. The path of the excitation light passes through a part of the pinhole disc which will divide the sample illumination into several hundred points. A microlens disc allows the light to be focused on the pinholes beforehand, thus reducing light loss. When the disc rotates (from 1500 rpm to 5000 rpm depending on the version), the spiral position of the pinholes allows the sample to be fully illuminated twelve times per revolution (figure 1). The fluorescence re-emitted by the sample is collected by the objective before passing through the pinhole disc. The pinholes allow only fluorescence rays perpendicular to the disc and coming from the focal plane of the objective to pass through. A dichroic mirror then reflects the fluorescence back to the camera. The resulting image has an axial resolution close to that obtained with conventional confocal laser scanning. The high rotation speed of the disc and the excitation at a relatively low light level allows very fast acquisitions and considerably limits photobleaching and phototoxicity.