Tissues were fixed in 10% neutral-buffered formalin and embedded in paraffin, and 5 μm sections were stained with hematoxylin and eosin (H&E) as described previously [52 (link)]. Immunohistochemistry was performed using a rabbit monoclonal anti-CD4 antibody (1:50; Epitomics, Burlingame/CA, USA) and a Benchmark XT automated slide staining system (Ventana, Tucson, AZ, USA) according to the manufacturer’s recommendations.
Microscopic findings were evaluated using a Zeiss Axioplan microscope (Zeiss, Oberkochen, Germany) with Olympus DP26 digital camera and CellSens Entry software (Olympus, Tokyo, Japan). The height of the mucosa was measured in triplicate, and a mean height value was calculated for each specimen. The number of CD4+ lymphocytes in the mucosa was counted in three different high power fields (HPF), and a mean value of CD4+ lymphocytes per HPF was calculated.
Microscopic findings were evaluated using a Zeiss Axioplan microscope (Zeiss, Oberkochen, Germany) with Olympus DP26 digital camera and CellSens Entry software (Olympus, Tokyo, Japan). The height of the mucosa was measured in triplicate, and a mean height value was calculated for each specimen. The number of CD4+ lymphocytes in the mucosa was counted in three different high power fields (HPF), and a mean value of CD4+ lymphocytes per HPF was calculated.
