Fig 1: The a2d1+pancreatic cancer cells own stem cell–like properties. (A) Immunofluorescent staining for a2d1 with mAb1B50-1 in living cells of indicated cell lines. Scale bars: 25 µm. (B) Flow cytometry analysis of a2d1+ fractions in indicated PDAC cell lines. (C) Relationship between a2d1+ cells stained by mAb1B50-1-FITC and those stained by PE-Cy5-labeled commercial anti-a2d1 antibody (Novus; Nb120-2864). (D) Representative micrographs demonstrating localization of a2d1 epitopes in indicated cells stained by 1B50-1-FITC and PE-Cy5-labeled commercial mouse anti-a2d1 monoclonal antibody (Nb120-2864). Scale bars: 25 µm. (E and F) Representative phase contrast micrographs (E) and histograms (F) showing primary (1°) and serially passaged (2°) spheroids formed by indicated subpopulations purified from indicated cell lines. Data represent mean ± standard deviation of 3 independent experiments. *Two-tailed Student t test. Scale bars: 100 µm. (G) Photographs showing dissected tumors formed by sorted a2d1+ and a2d1– cells from indicated sources. Bar = 1 cm. (H) Histology of tumors formed by a2d1+ cells sorted from indicated sources was compared with that of tumors formed by the respective parent cell lines as demonstrated by H&E staining. Scale bars: 50 µm. (I) Western blot analysis of expression of indicated molecules in FACS-purified a2d1+ and a2d1– fractions from indicated cell lines. (J) Flow cytometry analysis of percentage of a2d1+ fractions in parental cells (Parent), freshly FACS-purified a2d1+ subpopulations (Purified), and purified a2d1+ cells cultured in 10% fetal bovine serum for 2 weeks (Cultured) or engrafted into NOD/SCID mice (Tumor). (K) Percentage of a2d1+ fractions was analyzed by flow cytometry in FACS-sorted a2d1– subpopulations (Purified) and purified a2d1– cells cultured in 10% fetal bovine serum for 2 weeks (Cultured).