Nb100 98897

At the end of the study, the mini-pigs were euthanized by intraperitoneal injection of ketamine and xylazine (8 mg/kg and 10 mg/kg, respectively). Biopsies of upper thigh skin from the pigs were collected and processed for experiments as described^{8 (link),38 (link)}. Paraffin embedded pig skin tissues (Control or RTX treated) were cut and mounted on slides. Paraffin was removed and the tissues were rehydrated with different grades of ethyl alcohol, distilled water and PBS. Tissue sections were blocked in PBS, Triton X-100, and 5% normal goat serum. After washing with 1x PBS, the sections were exposed to rabbit polyclonal TRPV1 antibody (1:1000 dilution cat #NB-100-98897, Novus Biologicals), rabbit monoclonal PGP 9.5 antibody (1:500 dilution cat # ab-108986, Abcam, USA). Then, sections were treated with secondary antibody; goat anti rabbit IgG (1:1000; Thermofisher scientific, USA) and gently washed 3 times with PBS and cover slipped with Vectashield mounting media (Vector Laboratories). Colocalization of TRPV1 and PGP 9.5 in skin tissues was examined by confocal microscopy (Nikon, A1R Ti-E) or fluorescent microscopy (Nikon, Microphot FXA).