Sensory and motor evaluation of dogs with acute disc disease and spinal cord injury that undergo surgical decompression and canine adipose tissue-derived stem celltransplantation The adipose tissue was washed with phosphate buffered saline solution (PBS) (Gibco/Thermo Fisher Scientific, Grand Island, NY, USA) and tissue was digested using 1 mg/mL collagenase type I (Gibco/Thermo Fisher Scientific) for 30 minutes at 37°C, followed by 100 μm filter filtration (BD Biosciences Discovery Labware, Bedford, MA, USA). The cell suspension was centrifuged at 800 gfor 10 minutes and the contaminating erythrocytes were removed using a lysis buffer. Cells were washed, counted and plated at 1 × 105cells/cm2in 75 cm2culture flasks in Dulbecco's Modified Eagle Medium: Nutrient Mixture F-12 (DMEM-F12) (Gibco/Thermo Fisher Scientific) supplemented with 10% fetal bovine serum (FBS) (Sigma-Aldrich, Saint Louis, MO, USA) and 1% of antibiotics (Sigma-Aldrich).