High Throughput Vertical Flow Immunoassay Based On SERS Nanotags

With the rapid development of vitro diagnostic technology,the design of miniaturized and portable biochips has become a research hotspot nowadays.However,the current detection technology can only achieve low detection throughput,and different degrees of cross-reactions between samples exist.Especially in the context of epidemic prevention and control,medical resources are more difficult to access,and current detection methods are difficult to meet the need of simultaneous detection of multiple samples in hospitals.The vertical flow immunoassay technology has attracted extensive attention of scholars due to its advantages such as short detection time,high signal-to-noise ratio and strong multiplexing ability.The SERS nanotags based on surface enhanced Raman technology have multiple encoding capabilities,narrow emission spectrum,strong optical stability,and detection sensitivity can reach the level of single molecule.It is widely used in clinical testing,food safety,environmental protection and customs inspection.Based on it,this article designed a high-throughput vertical flow immunoassay chip,which combined the vertical flow immunoassay technology with SERS nanotags,and carried out a research on multiple parallel monitoring of inflammatory biomarker macromolecule and antibiotic small molecules.The main research content of this paper can be divided into the following three parts:(1)A detachable high-throughput vertical flow immunoassay chip was designed and fabricated,and the performance of the hydrophilic and hydrophobic region of the NC membrane was characterized.The selection of excitation wavelength was optimized,and the fluorescence background of NC film was the lowest at 785 nm.The choice of absorbent paper was optimized by introducing the vertical flow immunoassay common parameter:damkohler constant(Da)and pekelay constant(P_e),and compared the water absorption rate of the four kinds of absorbent paper,measured SERS signal strength in the vertical flow assay,and the cotton fiber paper was finally selected as the absorbent paper.The vertical-flow immunoassay chip provides a foundation for the subsequent multiple detection.(2)Single and multiple parallel detection of inflammatory factors were conducted based on four kinds of gold-core silver-shell SERS nanotags and high-throughput vertical flow immunoassay chip:the detection concentration gradient of four inflammatory factors(CRP,SAA,PCT,IL-6)ranged from 1 pg/m L to 10?g/m L,all covering the CUTOFF value.During the multiple parallel detection,the detection limit reached 30.2 fg/m L,22.4 fg/m L,18.6 fg/m L and 45.7 fg/m L respectively,which provides a new solution for clinical multi-sample mixed detection.(3)Single and multiple parallel detection of antibiotic small molecules were conducted based on three kinds of gold-core silver-shell SERS nanotags and high-throughput vertical flow immunoassay chip:the linear dynamic range of CAP,TC and LIN ranged from 1 pg/m L to 10?g/m L,covering a total of 6 orders of magnitude.In multiple parallel detection,the detection limit reached 0.495 pg/m L,0.039 pg/m L,0.207 pg/m L respectively,lower than their minimal inhibitory concentration(MIC)and predicted no effect concentration(PNEC).The optical mobile platform equipped on the small Raman spectrometer further realized the automatic measurement.