The invention discloses a three-dimensional super-resolution focusing method and device based on transparent medium pellet. The method comprises the steps of: emitting a radial polarized 
light beam to a micron-grade transparent medium pellet to focus to obtain a focused 
light spot at the exit end of the transparent medium pellet. The device orderly comprises a device for generating the radial polarized 
light beam, a micro-objective, the micron-grade transparent medium pellet and a 
cover glass for fixing the micron-grade transparent medium pellet, wherein the radial polarized 
light beam is primarily focused by the micro-objective and then further focused by the micron-grade transparent medium pellet to obtain the focused 
light spot. In the invention, a single light beam is used as a 
system working light beam, which saves 
system cost and increases practical applicability; simultaneously, the longitudinal axis length of the focused 
light spot is reduced to be less than 0.17 
lambda, the quality of the light beam is increased, and the transversal size of the focused light spot can be lower than the 
diffraction limit of the 
system; in particular, the volume of the focused light spot is small, which is favorable for actual application.