Figure 1

Multiparametric flow cytometry of PBMCs capturing uPAR/int β1 CTC subsets.

Breast cancer PBMCs were first sorted applying gating parameters to select for DAPI⁻ (4′, 6-diamidino-2-phenylindole)/EpCAM⁻/CD45⁻/CD44⁺/CD24⁻ cells. Cells were then subsequently sorted to obtain uPAR/int β1 subsets containing combinatorial expression of these markers. Antibodies used for flow cytometry and cell sorting were: anti-human CD45-APC-Cy7 (Biolegend, cat # 304015, 1:50 dilution), mouse anti-human EpCAM-PE CD326 (eBiosciences, cat # 12-9326-71, 1:40 dilution), anti-human CD24-PE ML5 (Biolegend, cat # 311106, 1:20 dilution), anti-human CD44-PE-Cy7 IM7 (Biolegend, cat # 103030, 1:20 dilution), mouse anti-human uPAR (CD87)-FITC (AbD Serotec cat # MCA2506FT, 1:10 dilution), anti-human int β1 (CD29)-ApC TS2/16 (Biolegend, cat # 3030008, 1:50 dilution). Cells were confirmed to be CTCs by performing RT-PCR, immunoflurescence and genotyping arrays. Representative images are shown.