Transport of the nonmetabolizable hexose analogue 3-O-methyl-D-glucose (3 OMG) was measured at 37°C, pH 7.4, in human polymorphonuclear leukocytes (PMNL) obtained from 10 ml of fresh venous blood. In the study, 0.05 mM of 3 OMG was equilibrated with a half time of about 10 sec, and the rate constant of entry was 0.074/sec in PMNL from a healthy subject (male, 38 yr). The coefficient of variation of values assayed on different days was 5.9% and intraassay variation was 2.4%. The mean 3 OMG transport rate in healthy subjects (N=-17; 14 males, 3 females; aged 25-38 yr) determined before lunch was about 9 fl/cell·sec. These results suggest that human PMNL may be a useful tool for the study of glucose transport in clinical investigations.