| Lateral flow biosensors?LFBs?have been widely used in the fields of medicine,food science and environmental monitoring because of their advantages of low-cost,user-friendly and fast-response without the involvement of specialized personnel or expensive machine.Most cancer biomarker detection methods are complex,time consuming and costly,which limits their use in early diagnosis of cancer.Thus,developing fast and sensitive Point-of-care?POC?analysis method to realize the detection of cancer biomarker with low abundance is of significant importance.However,the traditional LFBs have the disadvantages of low sensitivity and inaccurate quantification,hindering the further generalization for early biomarker diagnosis.Catalytic hairpin assembly?CHA?is an enzyme-free signal amplification method with low background,has been widely used in biosensor.Herein,in this thesis,to improve the detection sensitivity and quantitative detection ablity,two types of LFBs were designed for the rapid detection of microRNA-21?miRNA-21?.The detailed contents and achievements are described as follows:1.Catalytic hairpin assembly assisted signal amplification lateral flow biosensor for the rapid detection of miRNA-21.Due to the intrinsic feature of low abundance for miRNA-21,the traditional LFBs can't achieve the goal of accurate detection.To overcome this drawback in LFBs for nucleic acids detection without sacrificing its simplicity by integrating with CHA amplification.As a proof of concept,miRNA-21 was chose as the detection model and a signal amplification LFB was designed for the detection of miRNA-21.DNA hairpin probe 1?H1?was anchored on the gold nanoparticles?Au NPs?using Au-S chemistry at 5'end,while 3'end of H1 was modified with Bio.Due to the formation of the hairpin structure,biotin molecule on H1 probe was close to Au NPs and could not be captured by streptavidin?SA?on the cellulose membrane of LFB due to the steric hindrance effect.In the presence of miRNA-21 and the other hairpin probes?H2?,CHA amplification could be initiated on Au NPs to reuse the target,generating numerous double stranded DNA exposing biotin molecules outside.By monitoring the different shade of red bands on the strip,qualitative and quantitative analysis of target concentration could be easily realized by either naked-eye or reading the T line intensities with ImageJ software.The experimental results showed that the sensitivity of the signal amplification LFB designed in this experiment was nearly three orders of magnitude higher than that the traditional LFBs.Finally,the method was applied to detection miRNA-21 in tumor cells and serum samples,the feasibility and practicability were confirmed.2.Catalytic hairpin assembly assisted signal amplification lateral flow biosensor for SERS detection of miRNA-21.Improving the quantitative detection performance of conventional LFBs are of great importance to meet the demand of early clinical diagnosis of cancer.In this chapter,raman reporter-embedded gold-core silver-shell nanoparticles?Au@4-MBA@Ag NPs?were designed and constructed as SERS activity enhancement substrate,and CHA as a means of signal amplification for SERS quantitative detection of miRNA-21.Au@4-MBA@Ag NPs modified with biotinylated hairpin DNA were used as SERS labels,and the CHA reaction was used as a signal amplification strategy.As the CHA reaction went on,the hairpin DNA on the surface of the Au@4-MBA@Ag NPs SERS labels was opened,and can be captured by the interaction of Bio and SA.By monitoring the different shade of red bands on the T line,qualitative analysis could be easily realized;and the raman scattering intensity of 4-MBA on the T line was measured for the quantitative analysis of miRNA-21 with a detection limit of 8.4×10-14 mol/L. |
PDF Full Text Request