AF488 NHS ester 

AF488 NHS ester is an amine specific fluorescence probe (Em=525 nm). AF488 NHS ester reacts with sulfhydryl groups and amines in aqueous and biological samples then change their chemical structure and fluorescence properties after derivatization^[1].

Guide (The following is the experimental plan we recommend. This plan serves only as a reference guide. The specific operations should be adjusted according to your actual needs.)
1. AF488 NHS Ester Storage Solution^[1] Prepare 20 mM AF488 NHS ester using DMF.
2. Fluorescent labeling of AF488 NHS ester with IgG and solid-phase peptide library screening for anti-IgG^[2] Dissolve the freeze-dried IgG in a solution of 50 mM sodium phosphate, 20 mM sodium chloride, and pH 8.3 at a concentration of 5 g/L.
(2) Dissolve 1 mg of AF488 NHS ester in 100 μL of extra dry DMF, then add 1 mL of the solution from step (1), and rotate for incubation at room temperature for 1 hour.
(3) Use the Amicon Ultra 0.5-mL centrifugal filtration device, equipped with a 3-kDa MWCO filter, to collect the samples.
(4) The hexamers or tetramers were used to protect the library. It was washed three times with a 5× volume of precipitation resin in a solution of 50 mM sodium phosphate, 150 mM sodium chloride, and pH 7.4 (PBS) to achieve equilibrium.
(5) Dilute the IgG-AF488 with a 50 mM sodium phosphate, 150 mM sodium chloride, and 0.2% Tween (pH 7.4) solution to an final concentration of 1.3 mg/mL.
(6) Incubate (4) and (5) at 2-8°C for over night.
(7) Wash the resin beads with 50 mM sodium phosphate, 150 mM sodium chloride, 0.1% Tween 20, and pH 7.4 (PBS-T) solution.
(8) Deposit one bead in each well of a 96-well plate containing 40 μL PBS-T. Then, image using a fluorescence microscope at a magnification of 10x. Under 480 nm excitation, measure and screen for Alexa Fluor 488 fluorescence using a threshold of 510 nm emission intensity.

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

631.54

C₂₅H₁₇N₃O₁₃S₂

1374019-99-4

Solid

Orange to red

O=C(C1=C(C2=C(C=CC(N)=C3S(=O)(O)=O)C3=[O+]C4=C2C=CC(N)=C4S(=O)([O-])=O)C=CC(C(ON5C(CCC5=O)=O)=O)=C1)O

Room temperature in continental US; may vary elsewhere.

-20°C, sealed storage, away from moisture and light

*In solvent : -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture and light)

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture and light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

• [1]. Kendall CG, et al. Amine Analysis Using AlexaFluor 488 Succinimidyl Ester and Capillary Electrophoresis with Laser-Induced Fluorescence. J Anal Methods Chem. 2015;2015:368362.  [Content Brief]

  [2]. Lavoie RA, et al. Targeted Capture of Chinese Hamster Ovary Host Cell Proteins: Peptide Ligand Discovery. Int J Mol Sci. 2019 Apr 8;20(7):1729.  [Content Brief]