Janelia Fluor® 549, SE  (Synonyms: JF549, SE; JF549, NHS)

Janelia Fluor® 549, SE (JF549, SE) is a cell-permeable fluorescent dye with an NHS ester (succinimidyl ester (SE)) reactive group. NHS esters react with primary amines and are commonly used to conjugate dyes to proteins, antibodies, amine-modified oligonucleotides, and other molecules. Janelia Fluor® 549, SE also acts as a ligand for self-labeling tags such as HaloTag and SNAP-tag, and can be used in live-cell imaging studies (Ex/Em = 549 nm/571 nm)^[1].\n

Guidelines (The following are our recommended procedures; this is a guideline and should be modified according to your specific needs).
1. Dye Preparation
1.1 Preparation of Stock Solution
Prepare a 100 μM stock solution using anhydrous DMSO.
Note: It is recommended to aliquot the stock solution and store it at -20 ℃ or -80 ℃ protected from light.
1.2 Preparation of Working Solution
Dilute the stock solution with preheated serum-free cell culture medium or PBS to prepare a 10 nM working solution.
Note: Please adjust the working solution concentration according to your actual situation, and prepare it fresh before use.
2. Cell Staining (Halo-Tagged Cell Lines)
2.1 Culture adherent cells on sterile coverslips.
2.2 Remove the coverslips from the culture medium and aspirate excess medium.
2.3 Add 10 nM of the dye working solution, gently agitate to completely cover the cells, and incubate at 37°C for 30 minutes.
2.4 Aspirate the dye working solution and wash 2-3 times with PBS, 10 minutes each time.
2.5 After the third wash, replace the PBS with imaging medium (L-15 (Leibovitz) media containing serum (20%), glutamine (1%), and antibiotics (1%), and place a small amount of imaging medium into the microscope chamber.
2.6 Carefully transfer the coverslip to the cell culture chamber.
2.7 Properly assemble the cell culture chamber:
1) Use tweezers to place the coverslip on the circular part of the cell culture chamber base.
2) Close the lid of the cell culture chamber carefully, without squeezing.
3) Add an appropriate amount of imaging medium and check for leakage.
4) Place the assembled cell culture chamber in an incubator at 37°C.
5) Analyze the sample under a microscope.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

551.55

C₃₁H₂₅N₃O₇

1811539-32-8

Solid

Brown to black

549

505

O=C(C1=CC=C(C([O-])=O)C(C2=C3C=CC(N4CCC4)=CC3=[O+]C5=C2C=CC(N6CCC6)=C5)=C1)ON7C(CCC7=O)=O

Room temperature in continental US; may vary elsewhere.

-20°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

• [1]. Asaki Kobayashi, et al. A guide for Single Particle Tracking: from sample preparation and image acquisition to the analysis of individual trajectories. bioRxiv. April 04, 2025.