Blood samples of healthy adult humans (n = 40) and dogs (n = 30) grouped as males and females, were collected aseptically through appropriate measures. Thin blood smears were stained using Field stain ‘A’ and ‘B’. The PMNs were counted and observed for Barr bodies attached to them. Only the neutrophils with terminable lobes were examined and counted. The mean value ± SE of the Barr bodies was calculated and the difference between males and females of both species was analyzed through independent T-test. The mean ± SE values for Barr bodies was significantly higher (P ≤ 0.05) in human females (7.7 ± 0.63) as compared to their male counterparts (2.2 ± 0.38). However, statistically non-significant (P ≥ 0.05) result was noticed between female and male dogs being 1.8 ± 0.32 and 0.8 ± 0.24, respectively. In a nutshell, it is concluded that both the females of humans and dogs have a higher occurrence of Barr bodies on their PMNs as compared to their male counterparts. However, sex determination in humans through this cyto-diagnostic technique is quite certain and precise as compared to that in dogs.