An Ordered Binary Microbowl Array SERS Substrate And Its Application In Cancer Exosome Detection

Exosomes are the smallest type of extracellular vesicles,ranging in diameter from 40 nm to 160 nm.Exosomes play a crucial role in tumorigenesis,growth,metastasis,and drug resistance,and tumor-derived exosomes are consistent with the genetic content of tumor blasts.Therefore,exosomes and their transporters are increasingly regarded as a novel biomarker for cancer diagnosis.In this paper,a three-dimensional ordered binary microbowl array SERS substrate is proposed,combined with a microfluidic chip that can provide continuous flow conditions for highly reproducible SERS measurements and an improved SERS probe to explore high sensitivity to exosomes,high reproducibility and high stability detection,the main achievements of this paper are as follows:Firstly,polystyrene microspheres（PS）and silica microspheres（Si O₂）with hexagonal close packed structure were successively assembled on silicon wafer by interface assembly technology to prepare a double-layer microsphere template,and then based on this template,a polydimethylsiloxane（PDMS）-based ordered binary microbowl array was prepared.Subsequently,SERS active substrates with a large number of SERS"hot spots"were prepared by assembling a layer of uniform nano silver balls on the surface of the array through"three-phase interface assembly technology".Secondly,an improved gold-silver core-shell structure SERS probe was prepared.The preparation of the probe was performed by the sodium citrate reduction method,which made it have strong SERS activity and strong physical stability.In addition,the modification of PD-L1and EGFR aptamers achieved specific recognition of exosomes from breast cancer cells（MDA-MB-231）and lung cancer cells（A549）,respectively.Finally,a SERS microfluidic chip is constructed by embedding the substrate into the channel of the microfluidic chip by the method of bonding after oxygen plasma treatment,which has the advantages of high throughput,high sensitivity and low response time,while reducing the experimental operational complexity and cost.Based on the chip and the"sandwich"immune structure,the quantitative detection and screening of exosomes from different cancer cells was realized,which proved the versatility and feasibility of this scheme in the early diagnosis of different cancers.