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Ratiometric Fluorescent Biosensor for Hyaluronidase with Hyaluronan As Both Nanoparticle Scaffold and Substrate for Enzymatic Reaction
journal contribution
posted on 2014-09-08, 00:00 authored by Huafei Xie, Fang Zeng, Shuizhu WuHyaluronidases
(HAase) are involved in various physiological and
pathological processes and have been reported as urinary marker for
bladder cancer. In this study, a novel ratiometric fluorescent sensing
system based on both aggregation-induced emission (AIE) and aggregation-induced
quenching (ACQ) was developed to quantitatively assess hyaluronidase
level. First, a tetraphenylethylene derivative with positive charges
(TPE-2N+, typical AIE molecule) at both ends and an anthracene
derivative with positive charge at one end (AN-N+, typical
ACQ molecule) was synthesized. These two positively charged compounds
were then mixed with a negatively charged hyaluronan (HA), which induced
the aggregation of the compounds as well as the nanoparticles formation
as a result of electrostatic complexation, with TPE-2N+ acting as cross-linking agent. The aggregation also caused the efficient
quenching of the emission of AN-N+ due to ACQ effect, as
well as the fluorescence enhancement of TPE-2N+ due to
AIE effect. In the presence of HAase, the enzymatic reaction led to
the degradation of HA and triggered disassembly of the nanoparticles;
as a result, the emission of AN-N+ was restored and that
of TPE-2N+ was suppressed. This fluorescence variation
affords the system a robust ratiometric biosensor for HAase, and the
ratio of fluorescence intensity for AN-N+ (I414) to that for TPE-2N+ (I474) can be used as the sensing signal for detecting HAase
activity. In this system, hyaluronan serves not only as the scaffold
for nanoparticle formation but also as the substrate for enzymatic
reaction. This assay system is operable in aqueous media with very
low detection limit of 0.0017 U/mL and is capable of detecting HAase
in biological fluids such as serum and urine. This strategy may provide
a new and effective approach for developing other enzyme assays.
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ACQ effecthyaluronanhyaluronidase leveldetection limitchargeNanoparticle ScaffoldAIE moleculeTPEaggregationcompoundfluorescence intensityHAase activityfluorescence enhancementAIE effectassay systemenzyme assaysACQ moleculequenchingnovel ratiometricfluorescence variationbladder cancerEnzymatic ReactionHyaluronidasesnanoparticles formationemissionratiometric biosensornanoparticle formation
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