The size of a virion is a key criterion to its proper classification and may have implications in many practical aspects. Size determinations by thin section electron microscopy often result in length aberrations of more than 10% because of a number of preparative and instrumental inaccuracies, e.g. specimen shrinkage or swelling and unreliable calibration. Using adenovirus type 2 as an isometric size marker for internal calibration, we have determined the diameters of mature and immature HIV-1 to be 110 to 128 and 132 to 146 nm, respectively. The marker had been used either as a purified particle suspension added to the HIV producing culture, or adenovirus had been propagated together with HIV by infecting HIV producing cells. Using well characterized isometric markers, e.g. an icosahedral virus, in thin section electron microscopy appears to be a suitable technique for viral size determinations.