X-linked hypophosphatemic (Hyp) rickets is characterized by short stature, rickets, and bone abnormalities. Biochemically, hypophosphatemia and decreased renal reabsorption of phosphate are the hallmark of the disorder. Mutation of the PEX gene has been linked to human and murine Hyp rickets. Our study showed that phenotypical changes of this disease could be detected in 6-wk-old mice, but not in 2-wk-old mice. Therefore, we developed a PCR method to identify Hyp mice by detecting a lack of the 3' region of the PEX gene. Serum inorganic phosphate (Pi) levels were decreased, whereas alkaline phosphatase activity was increased in 2- and 6-wk-old Hyp mice. Northern blot showed that renal Na(+)-Pi transporter mRNA levels were decreased by 2.1-fold (1.47 +/- 0.21 densitometric units for normals; 0.68 +/- 1.43 for Hyp mice; p < 0.040) in 2-wk-old Hyp mice and by 1.7-fold (2.41 +/- 0.42 for normals; 1.44 +/- 0.33 for Hyp mice; p < 0.027) in 6-wk-old mice. Western blot showed that levels of immunoreactive renal Na(+)-Pi transporter protein were decreased by 4.5-fold (0.90 +/- 0.10 for normals; 0.22 +/- 0.08 for Hyp mice; p < 0.001) in 2-wk-old Hyp mice; and by 4.9-fold (1.47 +/- 0.19 for normals; 0.30 +/- 0.09 for Hyp mice; p < 0.0001) in 6-wk-old Hyp mice. In addition, levels of Na(+)-Pi transporter mRNA and protein were increased between 2- and 6-wk-old normal mice, but not in Hyp mice. This study demonstrates an easy assay to detect Hyp mutation and characterizes the defect during ontogeny of the Na(+)-Pi transporter in Hyp mice.