Cd90 thy 1

Immunofluorescence staining was performed 43, 51. Briefly, cells were seeded in 24‐well plates overnight, fixed with 4% paraformaldehyde, permeabilized with 1% NP‐40 and blocked with 10% donkey serum (Jackson Immuno‐Research Laboratories, West Grove, PA, USA), followed by incubating with CD73, CD105/endoglin, CD90/Thy‐1, CD166/ALCAM, BMPR‐II, CD117/c‐kit or CD29/Integrin β1 antibody (Santa Cruz Biotechnology, Dallas, TX, USA) for 1 hr at room temperature, as previously reported 43, 56. After being washed, cells were incubated with Texas Red or FITC labelled secondary antibody (Jackson ImmunoResearch Laboratories) for 30 min. Cell nuclei were counterstained with DAPI. Stains without primary antibodies were used as negative controls. Fluorescence images were recorded under an inverted fluorescence microscope.

Cell cultures were fixed with 4% of paraformaldehyde solution in phosphate buffer for 10 min, followed by a phosphate buffered saline (PBS) wash, pH 7.2, and blocked for 5 minutes in Power Block (Biogenex San Ramon, USA). The primary antibodies were incubated at 4°C overnight, and the secondary antibody was incubated for 1 hour at room temperature. The following primary antibodies were used: mouse anti-human vimentin (Chemicon, CBL202), CD90 Thy-1 (Santa Cruz, H-110) and rabbit anti-glucagon polyclonal (Linco, 4031). The secondary antibody was MULTILINK (Biogenex, HK268). Texas Red streptavidin (Vector, SA-5006) was added for 1 hour at room temperature, and nuclei were counterstained with Höescht staining. Imaging was performed using an epifluorescence microscope (Nikon, Eclipse E400).