Study On A Fluorescent Biosensor Based On Fluidic Magnetic Separation And Quantum Dots Labeling

Food safety incidents have occurred occasionally in recent years,which not only caused huge economic losses,but also seriously threatened public health.Foodborne pathogens are the main risk factors for food poisoning,thus,rapid screening of pathogens is the key to effectively prevent and control the outbreaks of foodborne diseases.The complex background of food samples often causes serious interference,and thus leads to false positive results.Besides,the concentration of pathogens is usually low,which makes currently available methods difficult to detect the pathogens directly.Therefore,it is of great significance to develop efficient separation and sensitive detection technologies of foodborne pathogens.This study aimed to develop a fluorescent biosensor for fast and ultrasensitive detection of foodborne pathogens and provide a pathogen bacteria detection platform for food safety.The proposed fluorescent biosensor consisted of immunomagnetic nanoparticles(MNPs),a coaxial capillary and circle ring high gradient magnetic fields for fluidic magnetic separation of bacteria,immune quantum dots(QDs)and an optical detection system for bacteria detection,and peristaltic pumps and a controller for fluidic control.First,the high gradient magnetic field was used to uniformly immobilize the MNPs in the channel of the coaxial capillary,and the target bacteria flowing through the channel were specifically captured by the monoclonal antibodies on the MNPs to form the MNP-bacteria complexes(magnetic bacteria).Then,the QDs modified with the polyclonal antibodies were used to react with the magnetic bacteria to form the MNP-bacteria-QD sandwich complexes(fluorescent bacteria).Finally,the optical detection system was used to measure the fluorescent intensity of the QDs on the fluorescent bacteria for quantitative detection of the target bacteria.To verify the feasibility of the proposed fluorescent biosensor for the detection of foodborne pathogens,E.coli 0157:H7 was used as research model to carry out concept proof and performance evaluation.The experimental results showed that:(1)The fluidic magnetic separation method based on the coaxial capillary and high gradient magnetic field could separate the target bacteria from a large volume(10 mL)of sample with an efficiency of up to 86%;(2)A good linear relationship between the fluorescence intensity(I)at the characteristic wavelength(615 nm)and the bacterial concentration(C)ranging from 101 to 105 CFU/mL was obtained and could be described as I = 48.18×lg(C)+132.15(R2=0.98);(3)The lower detection limit of the biosensor was 14 CFU/mL based on signal-to-noise ratio of 3;(4)When different concentrations of the E.coli 0157:H7 cells were spiked into the milk,the recoveries of the target bacteria were 95.92%-108.15%,and the detection time was less than 2 h;(5)When different volumes(10 mL and 1 mL)of sample with the same amount(about 10,000 bacteria)were tested,the detection results were basically the same,indicating that the biosensor could achieve specific separation and sensitive detection of the target bacteria in the large-volume samples.The proposed fluorescent biosensor could achieve fast and sensitive detection of E.coli 0157:H7 in a large volume(10 mL)of milk samples.By changing the antibodies or other biological identification materials,it can be extended for sensitive detection of other bacteria or pathogenic microorganisms,and immune quantum dots with different wavelengths can also be used to carry out simultaneous detection of multiple targets.