Mosaicism in blood varies with age, and cross-sectional studies indicate that for women, skewness of X-chromosomal mosaicism increases with age. This pattern could, however, also be due to less X-inactivation in more recent birth cohorts. Skewed X-chromosome inactivation was here measured longitudinally by the HUMARA assay in 67 septuagenarian and octogenarian women assessed at 2 time points, 10 years apart, and in 10 centenarian women assessed at 2 time points, 2–7 years apart. Skewed X-chromosome inactivation was also compared in 293 age-matched septuagenarian twins born in 1917–1923 and 1931–1937, and 212 centenarians born in 1895, 1905 and 1915. The longitudinal study of septuagenarians and octogenarians revealed that 16% (95% CI 7–29%) of the women developed skewed X-inactivation over a 10-year period. In the cross-sectional across-birth cohort study, the earlier-born septuagenarian (1917–1923) and centenarian women (1895) had a higher degree of skewness than the respective recent age-matched birth cohorts, which indicates that the women in the more recent cohorts, after the age of 70, had not only changed degree of skewness with age, they had also undergone less age-related hematopoietic sub-clone expansion. This may be a result of improved living conditions and better medical treatment in the more recent birth cohorts.