Research On Microfluidic Chip For BNP Fluorescence Immunoassay

Heart failure is one of the common chronic cardiovascular diseases in clinical practice,so the development of a portable microdevice for POCT of heart failure has an important applicable prospect.Brain natriuretic peptid(BNP)is a gold marker for early heart failure detection.Heart failure tests are expensive,misdiagnosed and time-consuming.Therefore,it is necessary to develop an instant detection portable microdevice for detecting heart failure marker BNP,to shorten the time of the detection process and improve the diagnosis rate of heart failure in emergency room.Currently,there are few microdevices mainly used for BNP detection in the market,and the development potential of heart failure real-time detection is huge.As a result,this paper has carried out the following research work:(1)The design method of BNP real-time detection microdevice structure is studied.Chip structures including injection zone,filtration zone,dissolution zone,mixing zone,detection zone and waste liquid recovery zone are designed.The microstructure of the detection unit was designed,and the influences of different microcolumns with the diameter of the circumferential circle of 0.1mm on the velocity and flow field in the detection unit were compared.The velocity field of the detection unit was simulated by using the microhydrodynamics module in the finite element simulation software Comsol,and the structure and size of the detection unit were determined.(2)Research on manufacturing methods of real-time detection devices was carried out.Firstly,a reasonable technological process was developed,and the silicon die manufacturing was completed by using lithography and dry etching processes.The channel structure of testing unit was hot-pressed on PMMA substrate with silicon die.The deviation between the channel depth and the design depth of the hot-pressing substrate was-0.08~0.57μm,and the deviation between the micro column height and the design height was-0.31~ 0.18μm.Finally,a reversible bonding method is proposed,which can be used to obtain the detection unit by reversible bonding of PMMA cover sheet and substrate.(3)The surface amination method of PMMA was studied,and the fixation of BNP capture antibody on the microchannel surface of the device was completed.The performance of real-time detection chip was tested by BNP test.Full spectrum scanning was conducted by X-ray electron spectrometer,and narrow spectrum scanning was conducted for nitrogen elements.It was known that the electron binding energy of N 1s was 398.7eV,which was consistent with the characteristic peak of N atoms in-NH2.Therefore,the surface group of PMMA after treatment was determined to be-NH2.In order to improve the sensitivity of the detection chip,the appropriate concentration of BNP antibody was selected.BNP concentration detection was realized by using double antibody sandwich method,and the corresponding calibration curve was determined.The theoretical detection limit of BNP is 5.06 ng/mL,and the detection results showed a linear range of 1~1000 ng/mL.