The expression of the large amino acid transporter, LAT1, was investigated in brain of adult Long-Evans rats. The LAT1 transcript was readily detected in brain microvessels and choroid plexus by reverse transcription polymerase chain reaction analysis using three different gene specific primer pairs. A polyclonal affinity purified antibody against the N-terminus of LAT1 was generated in chickens and used in immunoblot and immunocytochemical analyses of brain tissue sections of adult rats. On immunoblots, the antibody detected a peptide-inhibitable 45 kDa band in a rat brain microvessel membrane preparation. It also identified the same protein band in membrane preparations of different brain structures, as well as in heart and testis, whereas the protein was absent or only faintly detectable in muscle, kidney, and liver. In brain sections, the antibody intensely labeled the luminal and abluminal membranes of brain microvessel endothelial cells in all brain areas examined including cerebral cortex, cerebellum, hippocampus, and in gray and white matter regions. These results suggest that LAT1 is involved in transcellular transport and may play an important role in large, neutral amino acid transfer across the blood-brain barrier.