Design And Application Of Functional Nucleic Acid Based Lateral Flow Biosensors

Functional nucleic acid lateral flow biosensors were widely used in the field of rapid detection of food safety,because they integrated functional nanotechnology and functional nucleic acid technology.The ingenious integration has overcome the target range limitations,lack of sensitivity,poor thermal stability in traditional immune lateral flow biosensors,which provided a good choice and analysis platform for the rapid detection of target substances.However,there were still some aspects that need to be improved and optimized of the functional nucleic acid lateral flow biosensors,such as the lack of biotechnology foods and toxicology-related cancer biomarkers detection,the detection performance need to be further improved,the multiple inspections needs to be further optimized,and the quantitative equipment need to be further simplified.This paper will focus on the development of a series of novel functional nucleic acid lateral flow biosensors to solve the above problems.The main research contents were divided into the following four aspects:1.In this chapter,a "gold nanoparticle-antibody" functional probe was used to develop a universal nucleic acid lateral flow biosensor based on the "haptens tag-antibody" recognition system and "multiple-line" multiplex detection format,which can be used to detect tumor suppressor gene P16 methylation by employing proportion competitive quantitative PCR(PCQ-PCR).PCQ-PCR was based on the consistency of target ratio to obtain a variety of amplification products with universal tags in the middle and specific tags on the end."Multiple-line" multiplex detection format was based on the design of adding more test lines on a single carrier to enable simultaneous detection.This design was a general-purpose platform that can be applied to the detection of any other multiple target sequences through the adjustment of primers.2.In this chapter,a "gold nanoparticle-antibo dy-G-quadruplex" functional probe was used to develop a G-quadruplex nucleic acid lateral flow biosensor based on the "antigen-antibody" recognition system and "multiple-strip" multiplex detection format,which can be used to detect genetically modified food by employing tag-labeled multiplex LAMP(TM-LAMP).The signal enhancement technique was based on the deposition of specific red non-dissolved products from the reaction between AEC/H202 and G-quadruplex on gold nanoparticles,thereby increasing the signal intensity of the test line.The"mountain" shape design of "multi-strip" multiple detection form avoided the cross-reaction of multi-line interfaces and played an important role in improving the detection specificity.The sensitivity of this method has been improved an order of magnitude and the detection specificity was good as well.3.In this chapter,a "gold nanoparticle-DNA" functional probe was used to develop an artificial nucleic acid lateral flow biosensor based on the "artificial nucleic acid-normal nucleic acid" recognition system and "multiple-strip" multiplex detection format,which can be used to detect microRNAs by employing base stacking hybridization and exponential isothermal amplification(BSH-EXPAR).Peptide nucleic acid(PNA)has better specificity and affinity with DNA.Introducing PNA as probe in the development of nucleic acid lateral flow biosensor can significantly enhance detection specificity and accuracy.BSH-EXPAR reduced the interference of the background signal.The detection specificity and accuracy were significantly enhanced,and the single base difference of microRNA can be distinguished,which had important scientific significance and broad application prospects.4.In this chapter,a "gold nanoparticle-DNA" functional probe was used to develop a nucleic acid base mismatched lateral flow biosensor based on the "nucleic acid base mismatches" recognition system and "single-strip" detection format,which can be used to detect Hg(II)by employing cresol red dot and commercial apps in smart phones."Nucleic acid base mismatches" help that only one thymine-rich probe was immobilized on the detection line to capture mercury ions.Cresol red dot acted as reference color block to build self-made independent quantitative system,which can overcome the drawbacks of the traditional method.Commercial apps in smart phones were used to analyze test lines and reference color block for RGB.Finally,a "three-one" portable quantitative test assay,namely "a probe-a cresol red dot-a mobile phone",was provided to be an ultra-sensitive,practical,and efficient solution for safe and rapid detection in food.