Cyanine5 DBCO hexafluorophosphate (solution)

Guide (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs).
1. Preparation of solution preparation^[1][2]
1.1 Storage solution
Dissolve the Cyanine5 DBCO powder in DMSO (dimethyl sulfoxide) to create a stock solution, with a recommended concentration of 20 mM.
1.2 Preparation of working solution
Dilute the stock solution to 20 μM using PBS (Phosphate-Buffered Saline) or serum-free medium immediately before use.
Note: To enhance solubility, heat the tube to 37°C and use an ultrasonic bath if needed.


2. Cell Labeling (e.g. A549 cells)
2.1 Add 2 mL of growing seed cells at a density of 3×10⁴ (varies depending on the cells) to a 35 mm glass-bottomed petri dish.
2.2 Add Ac4ManNAz (50 μM, final concentration), and incubate for 3 days (forming azide groups on the cell surface).
2.3 Wash the cells twice with DPBS (pH 7.4).
2.4 Incubate the cells with Cyanine5 DBCO (20 μM, final concentration) at 37°C for 1 hour.
2.5 Wash the cells with DPBS (pH 7.4), and fix them with a combined fixative of formaldehyde and glutaraldehyde at 25°C for 15 minutes.
2.6 Wash the cells twice with DPBS (pH 7.4) and stain them with DAPI to label the cell nuclei.
2.7 Use a confocal laser scanning microscope to measure the fluorescence of Cyanine5 DBCO (Ex = 635 nm, Em = 650-700 nm).
Note:
1) After treatment with Ac4ManNAz, Cyanine5 DBCO can target the azide groups artificially introduced onto the cell surface for more than 3 days.
2) The degree of Cyanine5 DBCO labeling is dose-dependent.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only. Further protocols information, click here.