In myelodysplastic syndrome (MDS) as well as chronic myeloproliferative disorders (CMPD) others than chronic myeloid leukemia the frequency of pathognomonic genetic aberrations is very low and, therefore, X chromosome inactivation (XI) assays may help in assessing the clonality. To establish specific clonality criteria on XI, human androgen receptor assay (HUMARA) was performed on sorted myeloid and lymphoid peripheral blood cells of 21 healthy females. Clonality criteria 1 and 2 conferring at least 90% specificity were set based on the ranges and differences of XI number (XIN) describing the ratio of representation of the two alleles in as well as in between reactive myeloid and lymphoid compartments. Spiking experiments indicated that the test identifies clonality reliably when no more than 40-50% reactive cells are admixed. In the CMPD and MDS cases peripheral myeloid cells were monoclonal by one of the two criteria in 71-100%, whereas lymphoid cells in 28-75%. The results of HUMARA, available in 73% of the female patients, supported the clinicopathological data in 84% as well as proved pluripotent stem cell origin in 31-75% and 21% of CMPDs and MDS, respectively.