Trypsin ethylenediaminetetraacetic acid solution

Prostate cancer is the second leading cause of cancer-related death in men. Prostate cancer cells are good candidates to represent CTCs in peripheral blood.^{41 (link),62 (link)} Prostate cancer cell line LNCaP-C4–2 (passage #7, expressing green fluorescence protein (GFP) by lentiviral transduction) was grown in RPMI with 10% fetal bovine serum and 1% PenStrep (100 U/mL penicillin and 100 μg/mL streptomycin). Cells were grown in T-25 cm² culture flasks at 37 °C in a 5% CO₂ in air atmosphere until cells were ready for subculture. The morphology of the prostate cancer cells was observed before trypsinization (Figure 1). The cells were then detached from the flask with trypsin-ethylenediaminetetraacetic acid solution (Sigma-Aldrich). The LNCaP-C4–2 cells were trypsinized at 37 °C for 5 min, respectively. LNCaP-C4–2 cells were mixed with murine whole blood at about ~50 cells/mL. The white blood cells (WBCs) count in the murine whole blood is ~2–3 × 10⁶/mL; the lymphocytes count is ~1–2.5 × 10⁶/mL; and the platelets count is ~10⁹/mL.The size of blood cells including red blood cells (RBCs), WBCs, lymphocytes, and platelets are below 8–10 μm, while the prostate cancer cells are larger in size distributed mainly in the range of 10–15 μm according to previously published work.^{63 (link)}