The four main serotypes of Bordetella pertussis (1, 2, 3; 1, 2; 1, 3; 1) undergo spontaneous variation involving loss or gain of antigen 2 or antigen 3. By serial subculture from single colonies on charcoal-blood-agar medium, we have detected loss-mutations from type 1, 2, 3 to 1, 2 or 1, 3, and from type 1, 2 to type 1. Likewise we have found gain-mutations from type 1 to 1, 2 or 1, 3, and from 1, 2 to 1, 2, 3.These mutations apparently occur with a high frequency in some strains. Other strains have a lower mutation-rate and are more stable antigenically. We have not detected, by this method, either gain- or loss-mutations from the type 1, 3 strains that we have tested.These findings offer an explanation for the changes in serotype that occur during the course of a pertussis infection in the child and in the marmoset. They also constitute a warning on the possible antigenic instability of laboratory strains, especially relevant in the production, absorption and testing of diagnostic antisera and in the preparation of pertussis vaccine.