The CXCR4 expression profiles in the two types of BMSCs at two passages (P3 and P6) were assessed by immunofluorescence staining. Cells were collected and fixed in 4% paraformaldehyde at 4°C for 20 min. Following treatment with 0.5% Triton X-100 and 1% bovine serum albumin (Thermo Fisher Scientific, Inc.) for 1 h at 25°C, cells were incubated with rabbit anti-CXCR4 antibody (ab124824; 1:100; Abcam, Cambridge, MA, USA) overnight at 4°C. After washing with PBS, cells were incubated with Alexa Fluor® 488-conjugated anti-CXCR4 antibody (ab208128; Abcam) for 30 min at 37°C followed by DAPI (Sigma-Aldrich; Merck KGaA) staining for 2 min, to counterstain the cell nuclei. The sections were mounted and then observed under a Zeiss LSM 710 confocal microscope (Carl Zeiss AG, Oberkochen, Germany) and the intensity of the CXCR4-positive area were measured using Image-Pro Plus 6.0 software. The surface expression of CXCR4 was also analyzed using flow cytometric analysis as previously described (22 (link)). The cells were incubated with FITC-conjugated anti-CXCR4 antibody (Abcam).