By electron microscopy, particles of defective interfering Semliki Forest virus (DI SFV) had a mean diameter of 46.8 nm compared with 55.9 nm for standard virus particles, a decrease of 16%. The difference was confirmed by measurements of the two-dimensional projected areas of DI and standard virus particles. We examined nine different DI virus preparations produced by four to 13 undiluted passages in BHK cells and all were found to contain a majority of the smaller type of particle. Calculation of the absolute number of small particles showed that there were 130 particles per interfering unit measured by the inhibition of virus RNA synthesis. However, a more sensitive assay based on interference with virus protein synthesis gave a particle: interference ratio of 6.5.