Fitc conjugated anti cytokeratin ck3 6h5 antibody

Isolated cells were mounted, fixed on slides, and blocked for 5 min in 10% normal donkey serum. The cells were then stained with a phycoerythrin-conjugated anti-CD45 [5B-1] antibody (1:200; MACS Miltenyi Biotec). After staining, cells were permeabilized using 0.2% Triton X-100 in PBS containing 2 mM EDTA and 0.5% BSA. FITC-conjugated anti-cytokeratin [CK3-6H5] antibody (1:10; MACS Miltenyi Biotec) was used for cancer cell staining. For cell nuclei, cells were incubated with 4′,6-diamidino-2-phenylindole (DAPI). The sample was mounted with Dako Faramount aqueous mounting medium. Images were captured using a fluorescence microscope (Axio Scan.Z1, Zeiss, Jena, Germany). CTCs were defined as nucleated (DAPI positive) cells with cytokeratin (CK)-positive and CD45-negative staining.

Isolated cells were mounted and fixed on slides and were blocked for 5 min in 10% normal donkey serum (NDS). Hereafter cells were then stained with PE-conjugated anti-CD45 [5B-1] antibody (1:200; MACS Miltenyi Biotec). Cells were then permeabilized using 0.2% Triton X-100 in PBS containing 0.5% BSA and 2 mM EDTA followed by staining of the cells with FITC-conjugated anti-Cytokeratin [CK3-6H5] antibody (1:10; MACS Miltenyi Biotec). To stain the cell nuclei, cells were incubated in DAPI afterwards. The sample was mounted with Dako Faramount Aqueous Mounting Medium. Images were captured with a fluorescent microscope (Axio Scan.Z1, Zeiss). CTCs are defined as CK-positive, CD45-negative, and nucleated cells.

Isolated cells were mounted and fixed on slides and inhibited for 5 min in 10% normal donkey serum and subsequently stained with PE-conjugated anti-CD45 (5B-1) antibody (1:200; MACS Miltenyi Biotec, Bergisch Gladbach, Germany). Then, they were permeabilized using 0.2% Triton X-100 in PBS containing 0.5% BSA and 2 mM EDTA and stained with FITC-conjugated anti-cytokeratin (CK3-6H5) antibody (1:10; MACS Miltenyi Biotec, Bergisch Gladbach, Germany) and Alexa Fluor^® 647-conjugated anti-PD-L1 antibody [SP142] (1:100; Abcam, New Taipei City, Taiwan). To stain the cell nuclei, cells were incubated in DAPI afterwards. For mounting, Dako Faramount Aqueous Mounting Medium (Agilent, Santa Clara, CA, USA) was used. Images were captured using a fluorescent microscope (Axio Scan.Z1, Zeiss, Oberkochen, Germany). CTCs were categorized into CK-positive, CD45-negative, and nucleated.