The Na+-neutral amino acid (AA) transporter B0 functions for the uptake of luminal neutral AA across the gut apical membrane. The aim of this study was to quantify B0 mRNA expression along the jejunal crypt-villus axis in fed neonatal pigs. Six Yorkshire gilts were removed from sows at d 5 of age and fed a milk protein-based liquid formula to 14-16 d of age before tissue collection. Three epithelial cell fractions, representing cells from the upper villus, the middle villus and the crypt regions, were sequentially isolated, with cell viability of 92-95% assessed by trypan blue exclusion, along the crypt-villus axis from the entire jejunum by the distended sac method. The 443-bp porcine partial mRNA sequence of the neutral AA transporter B0 gene was obtained according to the conserved region in the human gene sequence. The partial porcine mRNA sequence had 86 and 85% homology with the known human and mouse sequences, respectively. Real time RT-PCR analyses (SmartCycler) using SYBR Green-I detection kit revealed no differences (P>0.05) in the B0 mRNA abundance between the upper and the middle villus cell fractions but a lower B0 mRNA level (P<0.05) in the crypt cells by using β-actin as the house-keeping control. These results suggest that the B0 gene expression is low in crypt cells and is increased with cell differentiation in the villus cells in the formula-fed neonate.