Fabrication Of Portable Biosensors And Their Application In Point-of-care Detection Of Nucleic Acids

Cancer poses great threat to health and has been one of the leading causes of death in many countries.A biomarker-based cancer diagnosis may significantly improve the early diagnosis and subsequent treatment,which can reduce cancer mortality.Nucleic acids(DNA and RNA),one kind of the major cancer biomarkers in cancer diagnosis have attracted much attention.In the past years,various traditional methods have been developed for nucleic acids detection such as microarray,polymerase chain reaction(PCR),in-suit hybridization.Although these methods can offer high accuracy and sensitivity detetions,most of them need expensive instruments,professional operators and complicated experimental process,which dramatically restrict their apllications in point-of-care(POC)settings.Thus,it is highly desirable to develop portable biosensors for POC detection of nucleic acids.This thesis is dedicated to the development of portable biosensor for nucleic acids detection at POC settings.The main contents are as follows:1.A cotton thread based nucleic acid biosensor with temperature-dependent pattern has been developed.Liquid wax was used to fabricate the temperature-dependent pattern to restrict the test zone in a narrow area,which could help to concentrate the detection signals and improve the sensitivity.With the accumulation of gold nanoparticles on the test zone,this biosensor enabled visual and quantitative detection of target DNA and a detection limit of 0.75 nM was achieved.In addition,the cotton thread based biosensor needed less sample than conventional lateral flow strip and the sample solution wicked faster at the cotton thread which could lead to a shorter detection time.This simple,low-cost and fast detection method held great potential to improve healthcare services in the developing regions.2.Inspired by biomineralization,DNA-Cu3(PO4)2 hybrid nanoflowers were prepared and exhibited high stability,high surface-to-volume ratio,good DNA encapsulation ability.Then a facile thread platform for miRNA detection was fabricated by employing DNA-Cu3(PO4)2 hybrid nanoflowers as captors,cotton thread as microfluidic channel and a personal glucose meter(PGM)as signal readout device.The fabricated biosensor could detect miRNA-21 quantitatively and a detection limit of 0.41 nM was achieved.Furthermore,miRNA in A549 cell lysate could also be detected without pretreatment.The developed portable biosensor showed great potential in biomedical and clinical applications.3.A three-dimensional(3D)DNA nanomachine for dual-modal detection of miRNA with fluorimeter and PGM was developed.The 3D DNA nanomachine was fabricated on magnetic beads(MBs),which was coated with amounts of hairpin structure of BHQ-H1-FAM(DNA tracks),activated by target miRNA-21(catalytic strand)and propelled by invertase-H2 conjugate(fuel strand)through strand displacement reaction.During these processes,the fluorescence of FAM on H1 was turned on(first signal)and the invertase on H2 was introduced to the surface of the MBs.After being separated by an external magnetic field,the invertase hydrolyzed the sucrose into glucose to generate the second signal,which was quantified by a PGM.The developed 3D DNA nanomachine showed high sensitivity and good specificity,the detection limits of 98 pM and 60 pM were obtained for fluorescence based assay and PGM based assay,respectively.Compared with the single-modal detection,the developed DNA nanomachine could achieve unique double signal readout and more reliable sensitive performance,which could satisfy different testing requirements both in laboratory and field and had potential applications in disease diagnostics and clinical applications.4.Inspired by self-cleaning property of titanium dioxide(TiO2),a renewable superwettable miRNA biochip(RSMB)was developed for specific and quantitative detection of miRNA-141.The hydrothermal synthesized TiO2 nanowires on the fluorine-doped tin oxide(FTO)coated glass substrate were employed as substrate for the fabrication of superhydrophilic-superhydrophobic pattern.Due to the extreme wettability difference between the superhydrophilic microwell and superhydrophobic substrate,the analytes could be concentrated and anchored onto the microwells from diluted solutions and achieved the sensitive detection of miRNA.In addition,the RSMB could be renewable efficiently via photodegrading the organics on the TiO2 substrate and consistent results could be obtained in several cycles.This work provided an alternative tool for developing the renewable biochips and showed potential applications in the biomedical diagnosis.