Cyanine5 DBCO (DBCO-Cy5) is a low-toxicity azide-reactive probe (near-infrared fluorescent dye) designed for imaging azide-labeled biomolecules via copper-free "click chemistry" [Ex=635 nm, Em=650-700 nm]. It exhibits no significant cytotoxicity in cells or animals and does not impact the physiological functions of non-target cells. Cyanine5 DBCO is suitable for labeling and tracking cells both in vivo and in vitro.

Guide (The following is our recommended solution. This solution is merely a guideline and should be modified according to your specific needs.): For solution preparation, 1) Stock Solution: Use DMSO as the solvent with a recommended concentration of 20 mM. Store in aliquots at -20°C or -80°C, shielded from light, and avoid repeated freeze-thaw cycles. 2) Working Solution: Dilute with PBS or serum-free medium to 20 μM (optimize for specific experiments), and use immediately while protecting from light. For cell labeling (e.g., A549 cells), 3) Seed 2 mL of growing cells at a density of 3×10^4 (cell-dependent) in a 35 mm glass-bottom culture dish. 4) Add Ac4ManNAz (50 μM, final concentration) and incubate for 3 days to produce azide groups on the cell surface. 5) Wash cells twice with DPBS (pH 7.4). 6) Incubate cells with Cyanine5 DBCO (20 μM, final concentration) at 37°C for 1 hour. 7) Rinse with DPBS (pH 7.4) and fix with a formaldehyde-glutaraldehyde solution at 25°C for 15 minutes. 8) Wash cells twice with DPBS (pH 7.4) and stain with DAPI to label nuclei. 9) Measure Cyanine5 DBCO fluorescence using a confocal laser scanning microscope (Ex=635 nm, Em=650-700 nm). Note: 1) After Ac4ManNAz treatment, Cyanine5 DBCO can target azide groups introduced on the cell surface for over 3 days. 2) Cyanine5 DBCO labeling is dose-dependent.