Detection Of Cancer Biomarkers By Microfluidics Coupling With Aggregation-Induced Emission

With continuous discovery of biomarkers in cancer diseases,the cancer biomarkers have been widely used in early screening,early diagnosis,curative monitoring and postoperative prognosis of cancers.Currently,rapid,high-throughput or multi-biomarkers joint detection cannot be achieved in one platform.In this regard,we propose a new method of aggregation-induced emission based on microfluidic chip to achieve multiple detection of cancer biomarkers.The main results are as follows:An antibody array construction method based on surface wettability-guided assembly technology has been developed to achieve simultaneous detection of multiple cancer biomarkers.We take advantage of the hydrophobic property of polydimethylsiloxane(PDMS)which remains on the slide after reversible bonding between PDMS and glass,and quickly construct hydrophobic region on the slide,while hydrophilic region forms microarray.Antibodies of different cancer biomarkers are immobilized in the hydrophilic pattern region of the slide and then incubated with the antigens and the horseradish peroxidase-labeled antibody.We immobilize antibodies of different cancer biomarkers in the hydrophilic lattice region of the slide.When the tyrosine functionalized tetrastyrene(TPE-Tyr)molecule is dissolved in an aqueous medium,the fluorescence emission is negligible.However,after the addition of horseradish peroxidase and hydrogen peroxide,the tyrosine in TPE-Tyr is effectively cross-linked by horseradish peroxidase-catalyzed oxidative coupling,thereby achieving aggregation-induced emission.In our design,each chip can simultaneously detect 9 different cancer biomarkers or the same biomarker in different populations.In the single detection of CEA as an example,the fluorescence intensity showed a good linear relationship with the antigen concentration in the range of5 ng/mL-100 ng/mL(R~2=0.9969),and the detection limit was 5 ng/mL.This chip provides a simple,efficient platform for multiple detection of cancer biomarkers.A microfluidic microwells array chip is constructed to achieve multiple cancer biomarkers detection.Arrayed microwells are constructed on the basis of PDMS and various antibodies are immobilized in the microwells.Then,the sample to be tested are delivered by microfluidic microchannels and are incubated with the biotin-labeled antibody,hereafter the unbound antibody are washed away.After that it is incubated with a mixture of streptavidin and biotinylated horseradish peroxidase.When biotinylated tetrastyrene(TPE-Biotin)is dissolved in a mixed solution of dimethyl sulfoxide and water,the fluorescence is negligible.But when streptavidin is present,TPE-Biotin molecules bind to the surface of streptavidin and initiate the aggregation reaction,and the fluorescence emission is remarkably enhanced.We utilize a multi-stage amplification system in the Elisa detection system to aggregate TPE-Biotin molecules and estimate the concentration of the antigen to be measured by fluorescence intensity.In this work,analysis of three different biomarkers are performed simultaneously in each chip as an example.In the single detection of CEA,the fluorescence intensity is positively correlated with the antigen concentration in the range of 5 ng/mL-500 ng/mL.The linear relationship between relative fluorescence intensity and antigen concentration is good in the range of 5ng/mL-100 ng/mL(R~2=0.9568)The chip has the advantage of enabling hemofiltration,automated solutions delivery and multiple biomarkers detection capability on the same device.