Cd45 pe antibody

To quantify analysis-disrupting white blood cell (WBC) contamination in tumor cell isolates, CD45+ cell count after manual and automated IMS was determined by flow cytometry. Therefore, we established a double staining protocol for leukocytes using a CD45-PE antibody targeting the specific surface protein of WBCs (1:50 dilution; Miltenyi Biotec, Bergisch Gladbach, DE, Germany) and nuclear stain RedDot™ (Biotium, Fremont, CA, USA). Wash buffers were set up in the IsoMAG IMS unit and 150 µL EpCAM beads were added to 7.5 mL whole blood before initiating the automated protocol. Manual enrichment was performed in parallel. The final samples were separated in a magnet separator, the supernatant was discarded and the bead cell pellet was stained with the above-mentioned solutions. Dual-positive cells [CD45+/RedDot+] were analyzed by FACS using the BD Accuri C6 flow cytometer. Cell counts after IMS were measured in four samples of different healthy donors and the average WBC contamination with standard deviation (SD) was calculated.