Sperry earned a bachelor's degree in English literature and a master's degree in psychology from Oberlin (Ohio) College and a doctorate in zoology from the University of Chicago in 1941. He then became an associate of Karl Lashley, first at Harvard University and then at the Yerkes Laboratories of Primate Biology in Orange Park, Fla. In 1946 he joined the faculty of the University of Chicago and in 1954 moved to the California Institute of Technology as Hixon professor of psychobiology.

Sperry's early research was on the regeneration of nerve fibres. He eventually became interested in brain function and undertook research on animals and then on human epileptics whose brains had been “split”—i.e., in whom the thick cable of nerves (the corpus callosum) connecting the right and left cerebral hemispheres had been severed. His studies demonstrated that the left side of the brain is normally dominant for analytical and verbal tasks, while the right hemisphere assumes dominance in spatial tasks, music, and certain other areas. The surgical and experimental techniques Sperry developed from the late 1940s laid the groundwork for much more specialized explorations of the mental functions carried out in different areas of the brain.