Ultrasensitive off-on-off fluorescent nanosensor for protamine and trypsin detection based on inner-filter effect between N,S-CDs and gold nanoparticles

Herein we develop a novel "off-on-off" fluorescent nanosensor for the ultra-sensitive detection of protamine and trypsin based on the as-prepared nitrogen and sulphur doped carbon dots (N,S-CDs) and gold nanoparticles (AuNPs). N,S-CDs were treated as fluorophore, while AuNPs were treated as fluorescence absorber. By experimental studies, the fluorescence quenching mechanism of N,S-CDs by AuNPs was inner-filter effect (IFE). Upon addition of protamine to the mixed aqueous of N,S-CDs/AuNPs, the electrostatic interaction and Au-N bond between protamine and AuNPs resulted in AuNPs aggregation, and the quenched fluorescence of N,S-CDs could be recovered. Protamine is rich in arginine and can be specifically hydrolyzed by trypsin, resulting in the deaggregation of AuNPs and further quenching the fluorescence of N,S-CDs. Thus, we can determine the concentration of protamine and trypsin by measuring fluorescence intensity of N,S-CDs. Under the optimized conditions, the fluorescence intensity showed a good linear relationship with protamine concentration of 20-260 ng/mL with the detection limit of 4.7 ng/mL. Meanwhile, quantitative determination of trypsin in a range from 10 to 140 ng/mL with the detection limit of 4.3 ng/mL can be achieved in this fluorescent nanosensor. This fluorescent nanosensor also showed excellent selectivity, and we successfully applied it for the detection of protamine and trypsin in human urine samples. Based on the above findings, we conclude that this novel "off-onoff" fluorescent nanosensor has great potential in clinical diagnosis.