*4.7. Immunohistochemistry*

Expression of LAT1 and LAT2 was also examined by immunohistochemistry (IHC) staining in paraffin-embedded placental tissue sections. A standard IHC staining procedure was performed as previously described [24]. Briefly, a series of deparaffinization was carried out with xylene and ethanol alcohol. Antigen retrieval was performed by boiling tissue slides with 0.01 mol/L citric buffer. Hydrogen peroxide was used to quench the endogenous peroxidase activity. After blocking, the sections were incubated with primary monoclonal antibodies specific against human LAT1 or LAT2 overnight at 4 ◦C. Corresponding biotinylate conjugated secondary antibodies and ABC staining system were subsequently used, according to the manufacturer0 s instructions. Slides stained with the same antibody were all processed at the same time. Stained slides were reviewed under an Olympus microscope, and images were captured by a digital camera with PictureFrame computer software.
