Analysis Method Of Single Cell Spectroscopy And Imaging Based On Microfluidic Droplets

Single-cell analysis is of great significance in understanding the activities of living organisms,disease diagnosis and treatment,and drug design.Microfluidic droplets provide an important analytical platform for single-cell studies due to their advantages of easy manipulation,small size and high throughput.However,the current single cell analysis technology based on microdroplet is still limited and challenging in detection methods,which is worth exploring continuously.The purpose of this paper is to develop single-cell analysis techniques combining microdroplet systems,including microdroplet-Surface enhanced Raman spectroscopy(SERS)and microdroplet-four-wave mixing(FWM)imaging.These two methods can be powerful supporting tools for single cell analysis.The main research is as follows:1.A high-throughput,two-channel single-cell analysis method was designed based on Plasma-SERS technology for the detection of sialic acid(SA)content on single cells.Multifunctional plasma nanoprobe was obtained by modifying 4-mercaptobenzene boric acid(MPBA)on silver nanoparticles(AgNPs).Due to the characteristics of AgNPs with strong energy loss,the nanoprobe is a high contrast indicator under strong field imaging and has a strong SERS enhanced capability for MPBA.The cells co-incubated with the plasma nanoprobe were wrapped in droplets and then redistributed in the detection chip with the chamber array.Through plasma imaging and image processing,the high-precision distribution of SA on the surface of individual cells was obtained.At the same time,SA expression levels of different cell lines(MCF-7,HepG2,SGC and BNL.CL2)were compared based on the changes of SERS spectrum.It was found that optical imaging and SERS spectrum were complementary to each other,and the differences of SA expression of four cells were successfully distinguished at the level of single cell.Therefore,the single cell analysis technique proposed in this study can provide an effective reference for the detection of single cell,single molecule and other biological fields.2.In order to improve the analysis efficiency,we first proposed a high-throughput single cell analysis technology based on the combination of droplet array and plasma enhanced four-wave mixing(PE-FWM)imaging on the basis of the above mentioned microdroplet-SERS platform.Metal nanoparticles are highly identifiable in PE-FWM imaging due to the multifrequency selection of their plasmas through resonance.This study simply modifies metal nanoparticles with capture molecules to form specific nanoprobes that specifically recognize receptors on the cell surface.Finally,the heterogeneity of cell surface receptors was analyzed by combining the high-throughput characteristics of microdroplet array with the automatic image processing.The two biomarkers expressed by the three cell lines were successfully distinguished at the level of single cell.In addition,the pe-fwm imaging platform proposed in this study has a high imaging contrast and is superior to bright field,dark field and fluorescence imaging,providing a good method for drug metabolism,biological imaging and other fields.