Abstract: Horseradish peroxidase (HRP) is a glycoprotein containing iron ions derived from horseradish that can be used for biochemical analysis, and catalyze the phenol in the presence of hydrogen peroxide. HRP can be combined with many substances to improve its performance and application. This work used a one-step hydrothermal method to easily synthesize fluorine-doped fluorescent carbon dots (F-CDs) with an average size of 2 nm. The as-synthesized F-CDs had obvious excitation light dependence, showing yellow-green fluorescence under ultraviolet light, and concentration dependence. The relative quantum yield was calculated to be as high as 45.6% with the quinoline sulfate as a reference. The results indicated that F-CDs have better fluorescence imaging performance in Hela cells, and F-CDs can form stable complexes (F-CDs/HRP) with HRP. The F-CDs/HRP complex not only preserved the ability of the enzyme to oxidize 2,2′-diazide-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), but also retained the excellent fluorescence performance of F-CDs. Moreover, the added hydrogen peroxide concentration will affect the ability of F-CDs/HRP to catalyze the oxidation of ABTS. At the temperature as low as 4 ℃, HRP still kept the activity to catalyze the oxidation of ABTS. Therefore, the catalytic activity of HRP in the obtained F-CDs/HRP could be analyzed qualitatively by the degree of chromogenic reaction. Therefore, the F-CDs/HRP complex has potential for application in immunofluorescence analysis.

Key words: horseradish peroxidase, carbon dots, fluorescence, immunofluorescence