Kinetic analysis of γ-glutamyltransferase reaction process for measuring activity via an integration strategy at low concentrations of γ-glutamyl p-nitroaniline

At 0.12 mmol/L γ-glutamyl p-nitroaniline(GGPNA),an improved integrated method was developed for kinetic analysis of γ-glutamyltransferase(GGT) reaction process and the integration with the classical initial rate method to measure serum GGT.For the improved integrated method,an integrated rate equation,which used the predictor variable of reaction time and considered inhibitions by both GGPNA and products,was nonlinearly fit to GGT reaction processes.For the integration strategy,classical initial rates were estimated when GGPNA consumption percentages were below 50%;otherwise,maximal reaction rates of GGT were estimated by the improved integrated method and converted into initial rates according to the differential rate equation at 0.11 mmol/L GGPNA.The inte-gration strategy was validated using optimized GGT kinetic parameters and 10-s intervals to record reaction curves within 8.0 min.By the integration strategy,there was a linear response from 0.9 to 32.0 U/L GGT,coefficients of variation were below 3.5%for GGT from 8.0 to 32.0 U/L(n=5) ,and GGT activities in clinical sera responded linearly to their classical initial rates at 2.00 mmol/L GGPNA with an expected slope.Therefore,the integration strategy was successful in measuring GGT at 0.12 mmol/L GGPNA.

Self-assembled all-inclusive organic-inorganic nanoparticles enable cascade reaction for the detection of glucose

Traditional colorimetric glucose biosensor generally involves complex assay procedures.Free labile enzymes and peroxidase substrates are used separately for triggering a chromogenic reaction.These limits result in inferior enzyme stability and defective enzymatic catalytic efficiency,making it hard to routinely utilize them for the direct and fast test of glucose.In this work,we provide an all-inclusive substrates/enzymes nanoparticle employed 3,3’5,5’-tetramethylbenzidine(TMB) as chromogenic substrates and glucose oxidase(GOx)/horseradish peroxidase(HRP) as signal amplifier enzymes(TMB-GH NPs) by the molecule self-assembly technique.The "all-inclusive" nanoparticles can realize the tandem colorimetric reactions,and the oxidation product of TMB(ox-TMB) exhibits a strong NIR laserdrive n photothermal effect,thus allowing qua ntitative photothermal detection of glucose.Owing to the restriction of the molecular motion of GOx,HRP,and TMB,the distance of mass transfer between substrates was s hortened largely,leading to improved catalytic activity for glucose.Overall,our strategy will simplify the analysis procedure,furthermore,these integrated nanoparticles not only display higher stability and activity than that of the free GOx/HRP system and possesses an excellent performance for colorimetric and photothermal bioassay of glucose simultaneously.We believe that this unique technique will give good inspirations to develop simple and precise methods for bioassay.