For CD26+ and chemokine-like receptor-1+ (CMKLR1+) MSC sorting, MSCs were digested into single-cell suspensions and separately incubated with an anti-human CMKLR1 PE-conjugated antibody (R&D, USA) or anti-human CD26 FITC-conjugated antibody (BD, USA) for 30 min. After washing with phosphate-buffered saline (PBS) three times, CD26+ MSCs or CMKLR+ MSCs were isolated with a BD Influx cell sorter for later experiments according to the manufacturer’s instructions. MSCs incubated with PE- or FITC-conjugated IgG antibodies were used as negative controls.
To detect the positive rates of CD26 and CMKLR1 expression, MSCs were then incubated with an anti-human CMKLR1 PE-conjugated antibody (R&D, USA) and anti-human CD26 FITC-conjugated antibody (BD, USA) as described above and detected using a BD Influx cell sorter.
To detect cell markers of MSCs, MSCs were incubated with anti-human CD29 PE-conjugated, anti-human CD44 FITC-conjugated, anti-human CD105 FITC-conjugated, anti-human CD14 FITC-conjugated, anti-human CD34 PE, and anti-human CD45 PE-conjugated antibodies (all from BD Pharmingen, USA) as described above and then detected using a BD Influx cell sorter.
To detect the positive rates of CD26 and CMKLR1 expression, MSCs were then incubated with an anti-human CMKLR1 PE-conjugated antibody (R&D, USA) and anti-human CD26 FITC-conjugated antibody (BD, USA) as described above and detected using a BD Influx cell sorter.
To detect cell markers of MSCs, MSCs were incubated with anti-human CD29 PE-conjugated, anti-human CD44 FITC-conjugated, anti-human CD105 FITC-conjugated, anti-human CD14 FITC-conjugated, anti-human CD34 PE, and anti-human CD45 PE-conjugated antibodies (all from BD Pharmingen, USA) as described above and then detected using a BD Influx cell sorter.