Quantum Dot-Aptamer Fluorescence Sensors For Rapid Detection Of Kanamycin And Tobramycin Residues

In animal husbandry and aquaculture,the unreasonable use of kanamycin and tobramycin causes them to remain in animal-originated foods,causing health problems after being eaten by the human body.Therefore,it is imperative to develop a simple and rapid method suitable for the specific detection of kanamycin and tobramycin residues on-site for strengthening market supervision.Aptamer is a kind of short single strand DNA or RNA with high affinity for specific targets.Quantum dots are a kind of semiconductor nanocrystals,which can emit multicolor fluorescence when excited at single wavelength.If they are bound as the biosensor that can be selectively labelled to specific targets,their fluorescent signals can be used to rapidly detect kanamycin and tobramycin residues.These are studied in this thesis,and the main results are as follows:（1）A method was developed for the synthesis of water-soluble CdTe quantum dots using thioglycolic acid as a stabilizer.The optimized synthesis conditions are as follows.The reaction time is 2 h,the reaction temperature 95^°C,the pH value 10,and the volume of thioglycolic acid 200μL.The synthesized quantum dots are spheres with a particle size of about 3.4 nm,and have good dispersibility.Carboxylated Fe₃O₄ nanomagnetic beads were prepared by a hydrothermal method.The magnetic beads are spheres with a particle size of about 340 nm and good dispersibility.（2）The methods were developed for the preparation of quantum dot-aptamer complex and complementary aptamer-nanomagnetic bead complex.The optimized reaction conditions are as follows.The molar ratio of quantum dot to aptamer is 1:6.8.The reaction temperature for the coupling of nanomagnetic beads and complementary aptamers is 25^°C,their reaction time is 3 h,and the amount of the complementary aptamers that can be coupled to the nanomagnetic beads are 0.237μmol/g.The quantum dot-aptamer complex and the complementary aptamer-nanomagnetic bead complex were successfully prepared by use of the developed methods.The quantum dot-aptamer-complementary aptamer-nanomagnetic bead complex fluorescence sensor was successfully assembled with the quantum dot-aptamer complex and the complementary aptamer-nanomagnetic bead complex by base pairing interaction.The signal of the sensor can be changed with the concentration of kanamycin.（3）A new method based on a quantum dot-aptamer fluorescence sensor was developed for the simultaneous detection of kanamycin and tobramycin residues.The optimized parameters of the detection system are as follows.The concentrations of the quantum dot-aptamer-complementary aptamer-nanomagnetic bead complexes of kanamycin and tobramycin reagent are 0.4 and 0.5 g/L,the incubation temperature is37^°C,and the incubation time is 30 min.Under the optimal conditions,the fluorescence intensity of the sensors are linearly enhanced with both kanamycin and tobramycin concentration within the range of 1-20μg/L.The detection limits are 0.11 and 0.21μg/L,respectively.The method was applied to the detection of kanamycin and tobramycin residues in milk.The recovery of spiked milk was between 95.3%-101.3%for kanamycin,and between 93.3%-97%for tobramycin.