5.7. Histological examination

The LCSs were harvested after 8 weeks of implantation and they were fixed with 4% formaldehyde. The fixed tissues were rinsed with tap water to remove the fixative for about 2 h. The LCSs were dehydrated in the graded ethanol and cleared in xylene using a tissue processor (Excelsior ES, Thermo Fisher Scientific, USA) and embedded into paraffin blocks sectionally using a paraffin embedding station (EG1150H; Leica, Germany). The paraffin blocks were cut into 5‐μm‐thick sections on a rotary microtome (RM2255; Leica, Germany). The paraffin sections were stained with hematoxylin and eosin (H&E) to identify the tissue structure inside the channels. For immunohistochemistry (IHC) staining of the sections, the following antibodies were used: rabbit polyclonal LYVE‐1 (1:100; NB100‐725, Novus Biologicals, CO, USA) as a lymphatic vessel marker, mouse monoclonal D2‐40 (1:100; ACR266B, Biocare Medical, CA, USA), which is a lymphatic endothelium marker, rabbit polyclonal CD31 (1:2000; ab182981, Abcam, UK), which is an endothelial cell marker. All histologic slides were acquired using a light microscope (Model BX40, Olympus, Japan).