Neutrophil functions and glucose metabolism are known to be impaired in glycogen storage disease (GSD) Ib patients. The uptake of nonmetabolizing glucose analogues into polymorphonuclear leukocytes (PMN) of GSD Ib patients was studied. 2-Deoxyglucose (2-DOG) and 3-O-methylglucose are transported across the cell membrane by facilitated diffusion mediated by the glucose transporter. Because 2-DOG is phosphorylated within the cell, its uptake rate reflects hexose transport as long as phosphorylation is not rate-limiting. These conditions prevail only at low 2-DOG concentrations. Transport of 5 µM DOG into GSD Ib patient PMN was found to be similar to controls (4.3 ± 0.5 and 4.65 ± 1.77 pmol/min x106, respectively). In contrast, 2-DOG uptake at high concentrations (2 mM) decreased by 70% in patient PMN compared with control cells (0.17 ± 0.06 and 0.51 ± 0.11 nmol/ min x 106, for patients and controls, respectively). Transport of 3-O-methylglucose (a glucose analogue that does not undergo intracellular phosphorylation) was not different in patient PMN compared with controls (1.86 ± 0.53 and 2.19 ± 0.30 nmol/min x 106, respectively). Hexose monophosphate shunt activity in PMN of GSD Ib patients at a glucose concentration of 2 mM was 43% of control values, whereas at 10 µM it was similar to controls. Taken together, these results suggest that the defect in glucose uptake and metabolism found in GSD Ib patient PMN is due to an impairment in hexose phosphorylation rather than in a reduction in the transmembrane glucose transport activity.