Figure 2 from PSA Secretion from Single Circulating Tumor Cells of Metastatic Castration-Naïve Prostate Cancer Patients

Workflow schematically illustrating the DLA processing to obtain viable CTCs. (1) DLA enrichment using the reduced enrichment reagent (RER) protocol; (2) staining using anti–PSMA-PE, anti–CD45-APC, and viability dye calcein AM; (3) sorting of calcein⁺ and CD45– CTCs enriched from the DLA; (4) seeding into the VyCAP nanowells; (5) imaging using a fluorescence inverted scanning microscope to visualize cells captured in the nanowells; (6) PSA secretion from single cells and PSA capture overnight (24 hours) using an anti–PSA-coated membrane and addition of anti–IgG-PE to visualize the imprint; (7) staining of the membrane for visualization of PSA secretion spots; and (8) correlation of PSA spots with cells with the help of imprint and the SPOT software to identify PSA-secreting and non-secreting PSMA⁺/PSMA⁻ CTCs.