Guidoni earned a doctorate in astrophysics from the University of Rome "La Sapienza" in 1978 and worked in the Italian Space Agency (ASI) as well as in the European Space Agency (ESA). One of his research projects was the Tethered Satellite System, which was part of the payload of the STS-46 mission. Guidoni trained as an alternate payload specialist for that mission and made his first spaceflight aboard the Space Shuttle "Columbia" STS-75 in 1996, which included the second flight of the TSS system (TSS-1R).