| In recent years,animal diseases such as mad cow disease,foot-and-mouth disease,swine streptococcosis,porcine reproductive and respiratory syndrome,avian flu,rabies and other animal diseases brought about a heavy blow to the global animal husbandry and had a huge impact on food safety and society economic life,which caused great attention on animal disease prevention and control and food safety.The early monitoring of animal disease markers is critical to the diagnosis and control of the disease.However,some traditional detecting methods of animal diseases are mostly sensory or optical assays have,mainly including virus neutralization test,latex agglutination test(LAT),enzyme-linked immunosorbent assay(ELISA),fluorescence analysis and polymerase chain reaction(PCR).Although most of the aforementioned methods are reliable and effective,they usually suffer from serious drawbacks,such as insensitive,time-consuming and expensive.Therefore,there is an urgent need to develop some simple,fast,cost-effective,sensitive and reliable new technologies or new methods to detect animal diseases.The work aim to develop new animal disease detection technology.From the perspective of improving the sensitivity and accuracy of detection technology we reasonably design and screen nano-material with unique function to construct three kinds of signal-amplified electrochemical luminescence sensors for ultra-sensitive detection of major disease markers.The main contents are as follows:1.Triple and multiple signal amplification of electrochemiluminescence immunosensor for ultra-sensitive detection of porcine pseudorabies virusIn this work,gold-graphene(Au-GN)nanocomposites and silicon-coated bipyridyl ruthenium nanoparticles(Si@Ru NPs)were prepared first.Au-GN nanocomposites were used as the electroactive matrix and Si@Ru NPs as the enhanced electrochemical luminescence signal tag.Relying on the interaction of antigen-antibody-secondary antibody and of biotin-streptavidin-biotin(B-SA-B),the triply-amplified biosensor could be fabricated and exhibited better analytical performance not only toward PrV monoclonal antibody with a linear detection range from 50 ng m L-1 to 1 pg m L-1 and a detection limit of 0.40 pg m L-1,but also toward PrV polyclonal antibody and actual serum samples when compared with enzyme-linked immunosorbent assay and fluorometry.Furthermore,multiply-amplified biosensors could be conveniently fabricated by controllably repeating the combination of B-Ru@SNPs and SA to form the “B-SA-B” structure.After repeating three times,the multiply-amplified biosensor stretched to the maximum of signal amplification and achieved a luminescence quantum efficiency about 23.1-fold higher than the triply-amplified biosensor.The tunable biosensor exhibits good stability,acceptable reproducibility and accuracy,suggesting its potential applications in food safe and clinical diagnostics.In addition,a three-step construction strategy of “stretch–stowage–growth” is proposed,which provides the idea for the construction of triple and multiple electrochemiluminescence biosensors.2.Near–infrared electrochemiluminesence biosensor for high sensitive detection of PRRSV based on signal amplification of carbon and PtAu nanotubesIn this study,the near-infrared CdTe/CdS quantum dots was prepared by regulating the reaction time.Porous platinum heterojunction nanotubes was prepared by tellurium nanowire template method.Based on carbon nanotubes carrier,the porous platinum heterojunction nanotubes,the host-guest recognition between ?-cyclodextrin and adamantane and “antibody-antigen-antibody” specific action,were fabricated a novel near–infrared electrochemi-luminesence immunosensor for ultrasensitive detection of PRRSV.Compared with the traditional veterinary laboratory diagnosis of PRRSV antigen like PCR and indirect immunofluorescent assay,the current method exhibits a higher sensitivity and wider linear range.The ECL peak intensities were proportional to the logarithmic value of the PRRSV dilution ratio over a range from1:103 to 1:106(19 ng/m L–19 pg/mL)with a detection limit of 10.8 pg/mL.In this work,the optical detection mode is extended from the visible region to the near infrared region,which avoids the spontaneous light interference of the blood sample or tissue.This work is helpful for achieving the detection of animal diseases with high sensitivity and low disturbance.3.Signal-amplified near-infrared ratiometric electrochemiluminescence aptasensor for detection of thrombin based on Graphene/Gold Nanorods/G-QuadruplexIn this study,the common model of thrombin was used to study the dual aptamers model for the detection of porcine epidemic diarrhea virus.The four-in-one graphene-gold rod-DNA enzyme composites were successfully prepared by using graphene,gold rod,hemin and G-quadruplex.The near-infrared CdTe/CdS quantum dots and luminol were used as negative potential signals and positive potential signals,respectively.A near-infrared ratio type electrochemical luminescence sensor with signal amplification were constructed based on the dual-aptamers sandwich mode.This strategy not only broadens the wavelength range of dual-signal ratiometric strategy from visible light to NIR,but also introduces signal amplification into ratiometric ECL sensors,which can meet the demand for high sensitivity and accuracy.The four-in-one intermediate media introduced in the sensor can induce electrochemical luminescence synergistic quenching of near-infrared quantum dots and synergistic electrochemiluminescence enhancement of luminol.Before and after the test,the ECL signal appears “shift” type of change.Quantitative detection was achieved by the correspondence between the ratio of the double signal intensity and the thrombin concentration.The value of ECLluminol/ECLQDs was linearly dependent on the logarithm of TB concentration in the range from 100 ng/mL to 0.5 pg/m L.The limit of detection(LOD)corresponding to a signal-to-noise ratio of 3 was estimated to be 4.2 fg/m L.Furthermore,the signal-amplified ratiometric ECL sensing strategy could serve as a universal“mode”for developing other amplified ratiometric ECL sensors by only using different synergetic nanocomposites,which sheds new light on the development of highly sensitive and accurate ratiometric ECL sensors... |
PDF Full Text Request