Fluorescence Biosensor For Cancer Marker Detection Based On ?-FeOOH Nanosheets Madiated Low Background And Enzyme-assisted Signal Amplification

The development of industry has not only brought about technological progress,but also caused many environmental problems.Ecological pollution is the chief culprit that destroys the environmental system and increases human,plant and animal diseases.In order to protect human health,early diagnosis and treatment of diseases are necessary.Many small biological molecules,as gene regulators,play an important role in a variety of biological processes and are necessary to maintain human metabolism.However,the abnormal expression of these small biological molecules in the human body poses a serious threat to human health.The current related clinical diagnostic techniques are highly sensitive and selective,but they require tedious sample preparation and pre-concentration procedures,expensive instruments and professionals.Therefore,the development of a detection method with high sensitivity,good selectivity,rapidity,simplicity and low cost has great application prospects.In this paper,using nano-functional materials to reduce the background value and combining signal amplification strategies,we designed and constructed ultra-sensitive and highly selective fluorescent biosensors,and applied them to the detection of biomolecules,providing a new way for the early diagnosis of some diseases.The main research contents are as follows:(1)A series of characterizations of the synthesized ?-FeOOH nanosheets proved the superior properties of the magnetic iron oxyhydroxide nanosheets.The experiment confirmed that ?-FeOOH nanosheets have different affinities for short oligonucleotide fragments and long ss DNA probes,and can be used as an effective quencher for DNA fluorescent probes.Moreover,the ss DNA probes adsorbed on the?-FeOOH nanosheets can be effectively protected to avoid digestion by the DNase I enzyme.Through the combination of ?-FeOOH nanosheets and DNase I enzyme-assisted target cycle signal amplification technology,a simple,sensitive and selective detection method for mi RNA-141 is proposed.The detection limit of this method for mi RNA-141 can reach 1 p M,which is 3 orders of magnitude lower than the non-amplified method.In addition,it is possible to distinguish between mi RNAs and mi RNA targets that do not match a single base.In addition,our method allows direct mi RNA analysis in complex biological samples without any sample pretreatment,which greatly simplifies the rapid detection procedures.(2)In this work,combined with DNaseI enzyme-assisted signal amplification and magnetic separation technology,the selective and highly sensitive detection of MUC1 was achieved.The sensitivity of this amplification strategy is 10 times higher than that of the traditional unamplified method.The fluorescence intensity is linearly related to the concentration of MUC1 in the range of 20 ng/m L to 200ng/m L,and the detection limit is 4 ng/m L.The results prove that the aptamer sensor has an absolute advantage in the specific detection of MUC1,and can be used for the detection of MUC1 in relatively complex samples.This design not only provides a new fluorescence sensing platform for the detection of MUC1,but also provides a new way for exploring 2D nanomaterials as biological applications.(3)In this work,a novel magnetically controlled two-dimensional nano-DNA fluorescence sensor for sensitive and selective detection of alkaline phosphatase(ALP)activity has been developed by combining the ? exonuclease(? exo)cleavage reaction and a ?-FeOOH nanosheet-based platform.A5'-phosphoryl-modified double-stranded DNA probe(5'-P-ds DNA)was designed as a substrate for ALP,and a 5'-dye-labeled single-stranded DNA/ ?-FeOOH system was designed as a detection unit.In the absence of ALP,5'-P-ds DNA was degraded by ? exo to yield mononucleotides.Thus,the fluorescence of the detection unit was quenched.However,in the presence of ALP,the 5'-phosphate functional group of5'-P-ds DNA was efficiently removed to form 5'-hydroxyl-ds DNA,which could no longer be degraded by ?exo.Then,the dye-labeled single-stranded DNA hybridized with the 5'-hydroxyl-ds DNA to form a triplex DNA(ts DNA),resulting in increased fluorescence due to the weak binding of ts DNA with the ?-FeOOH nanosheet.More importantly,a high signal-to-background ratio was obtained through magnetic separation.The detection limit for ALP was 0.02 m U/m L(3?/S),which is much lower than that previously reported for fluorescence methods.This work revealed the application of ?-FeOOH nanosheets in the design of magnetically controlled biosensors for biomolecular detection.