NBD-F is highly reactive and can label primary and secondary amines under mild conditions (1 min reaction at 60ºC in a weak basic solution). NBD-F is a pre-labeling compound for HPLC analysis of small molecules. NBD-labeled compounds are orange with a maximum wavelength at 470 nm.The excitation and emission of the derivatized compound are 470 nm and 530 nm, respectively.
1. To prepare sample solution, mix or dissolve a sample with 50 mM borate buffer (pH 8.0) containing 20 mM EDTA.
2. Mix 300 μl of the sample solution and 100 μl of 100 mM NBD-F/acetonitrile solution in a reaction vial.
3. Heat the vial at 60ºC for 1 minute and then cool it on an ice bath.
4. Add 400 μl of 50 mM HCl aqueous solution to the reaction mixture.
5. Use this mixture for HPLC analysis to determine NBD-labeled compounds.
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