Results of cross-neutralization tests of 4 isolates of contagious ecthyma (CE) virus and their antisera indicated that the isolates were neutralized to various degrees by the CE virus antisera. Cross-reactions among isolates were unilateral, but not bilateral, and therefore grouping of CE virus isolates by neutralization tests was not possible to obtain. Structural analysis of polypeptides from 11 isolates of CE virus by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the profiles obtained were similar to one another, except for differences observed in the molecular weight region of 37,000 to 44,000. On the basis of these differences, the isolates could be classified into 4 groups. The polypeptides which varied among the different isolates were shown to be in the surface component of the virion and appeared to be components of the surface tubules. Unilateral cross-reactions detected in cross-neutralization tests were found to correlate with the 37,000 to 44,000 dalton polypeptide grouping of the isolates.