Anti trop2

Immunocytochemistry was performed as described before 34 (link) using anti-OCT4 (sc-5279, Santa Cruz Biotechnology) and anti-TROP2 (BD Bioscience) antibodies. Cell nuclei were counterstained with DAPI.

CTCs were assessed by immunofluorescence staining. The following antibodies were used to identify CTCs and assess EPCAM and TROP-2 staining as indicated in the figure legends: Anti-CD45 (BioLegend Cat# 304018, RRID:AB_389336), Anti-CD34 (BioLegend Cat# 343508, RRID:AB_1877133), Anti-CD11B (BioLegend Cat# 101218, RRID:AB_389327), Anti-CD66B (BioLegend Cat# 305109, RRID:AB_2563170), Anti-Pan cytokeratin (BioLegend Cat# 628602, RRID:AB_439775 or Abcam Cat# ab49779, RRID:AB_869395), Anti-Androgen Receptor (Cell Signaling Technology Cat# 5153S, RRID:AB_10692774), Anti-EPCAM (Abcam Cat# ab112068, RRID:AB_10861805) or Anti-TROP-2 (BD Biosciences Cat# 940370, RRID:AB_2876239) and Hoechst 33342 (Thermo Fisher Scientific). Extracellular antibodies were stained at 4°C for 30 minutes. For intracellular and nuclear staining of cells (Fig. 2), cells were stained as described by Sperger and colleagues (30 (link)). For intracellular staining (Fig. 4), cells were permeabilized, stained, and washed with BD Perm/Wash. Images were taken with a 10x objective using Nikon Eclipse Ti-E with an ORCA-Flash 4.0 V2 Digital CMOS camera (Hamamatsu) and NIS-Elements AR Microscope Imaging Software (RRID:SCR_014329, Nikon Instruments). Images were background subtracted, and CTCs were determined by Hoechst-positive staining, cytokeratin⁺ and CD45⁻/CD34⁻/CD66b⁻.