Study Of Microfluidic Chip Integrated With Pretreatment And Sensing

The integration of all analytical processes on a single microfluidic chip is a fundamental feature of microfluidic analysis on the microchip.It also has the advantages of low consumption,high throughput,and easy integration.In recent years,much more attention has been paid to the introduction of optoelectronic sensing technology into the microfluidic analytical system which has shown great potential for solving the challenges of complex matrix interference,low target content,poor detection sensitivity and complicated operation that usually exist in biological sample detection.Microfluidic chip integrated with photoelectric sensors is gradually growing as a research hotspot for the rapid and highly sensitive detection of small biological molecules,proteins,exosomes,and cells in biological samples.It has become a new platform for biochemical analysis.In this thesis,the functional modules of pretreatment and sensing were proposed to be integrated on the microfluidic chip for the efficient detection of biological samples including target proteins in blood and microorganisms in urine.The integrated microfluidic chips and analytical system were designed and fabricated to detect biochemical targets by the proposed methods.The related research is of great value to the microfluidic analysis technology and photoelectric sensing technology,and has good practical prospects in the fields of life science,clinical medical diagnosis,and environmental monitoring.The main achievements of this work are as follows:(1)A microfluidic chip integrated with dielectrophoresis(DEP)separation and differential pulse voltammetry(DPV)sensing was designed and fabricated,and a highly sensitive and rapid method was established for the on-chip detection of the cancer marker prostate specific antigen(PSA)in whole blood samples.In this thesis,an electric-force coupled pretreatment module based on the dielectric and flow field effects was proposed to integrate on a microfluidic chip.The dielectric effect of interdigital array electrodes with different structures on blood cells and nano-signal amplification technology were investigated.The separation performance of the continuous DEP separation module based on the designed V-shaped interdigital array electrode and microchannels was investigated.It was shown that the separation efficiency of blood cells was up to 98%.The detection limit of PSA in whole blood was reduced by 2 orders of magnitude after effective separation of blood cells.An electrode for DPV sensing was integrated into the end of the chip using the synergistic effects of bio-selective recognition,nano-signal amplification,and enrichment by the nano-sensing interface.A sandwich sensing strategy based on dual-aptamers and nanoprobes was proposed to achieve the highly sensitive and selective detection of PSA in whole blood samples with a detection limit as low as 250 fg/m L and a linear range of 1 pg/m L to 10 ng/m L.The highly sensitive and rapid detection of PSA could be completed with only one injection,demonstrating that the proposed integrated chip had a good application prospect in the field of point-of-care testing(POCT)for clinical diagnosis.(2)A microfluidic fluorescence sensing chip integrated with DEP-magnetic bead enrichment dual-pretreatment was designed and fabricated,and an efficient method was established for the on-chip detection of multiple trace cancer markers,total and free prostate-specific antigen(t-PSA and f-PSA),in whole blood samples.The migration patterns of blood cells in dielectric field and immunomagnetic bead complexes in magnetic field were carefully studied.A DEP separation module based on the designed inverted V-shaped interdigital array electrode was constructed,and the separation efficiency of blood cells was up to 98%.An enrichment module based on immunomagnetic beads and microchannels was also constructed,and the enrichment efficiency of the targets was up to 694 times.An off-on fluorescence sensing method was then established for the on-chip detection of t-PSA and f-PSA in whole blood samples using the designed washing-free turn-on fluorescent nanoprobes,and the detection limits were 100 fg/m L and 500 fg/m L,respectively.The proposed integrated chip had a good application prospect in the field of detection of low abundance actual biological samples.(3)A microfluidic fluorescence detection chip integrated with multistage DEP separation and dialysis dual-pretreatment was designed and fabricated,and a new and efficient method for the on-chip detection of microbial in urine samples was established.A dialysis separation module based on spiral microchannels was designed to regulate the conductivity of urine by removing electrolytes from urine samples through the dialysis effect,and the conductivity of the urine samples was effectively reduced by this module.The multi-stage V-shaped interdigital array electrode was proposed to integrated on the chip for the manipulation of the microbial,by which the highly efficient separation of Candida albicans and Staphylococcus aureus could be achieved.The separation efficiency of Candida albicans and Staphylococcus aureus was 99%and 88%,respectively.The processing flux of urine by the proposed chip was increased by 36 times by the designed tandem separation mode of dialysis and multi-stage DEP that integrated on the chip.A quantitative method of the on-chip detection of Candida albicans and Staphylococcus aureus in urine was established using the washing-free nucleic acid dye SYBR Gold as the fluorescent reagent.The limits of the detection of Candida albicans and Staphylococcus aureus were as low as 10~1 CFU/m L.It was demonstrated that the proposed integrated chip could be a new technical approach and test platform for the efficient detection of microbial in actual biological samples.