Detection Of Exosomes Based On SERS For Diagonosis Of Hepatocellular Carcinoma

Hepatocellular carcinoma(HCC),one of the most common cancers across the world,especially poses a serious threat to public physical health in China.HCC has high incidence and serious deterioration yet without any obvious symptoms in the early stage.The majority of the patients with a clinical diagnosis of HCC are at terminal stage.Currently,the frequently-used diagnostic methods of HCC include imaging tests,liver biopsy and detection of serum alpha-fetoprotein(AFP).However,imaging tests,as the most commonly used method for the diagnosis of HCC in the clinic,is difficult to detect small hepatocellular carcinoma.Factors such as size or depth of liver tumor,to some extent,may lead to the false negative in liver biopsy.Serum AFP,though frequently-used in HCC diagnosis,shows poor sensitivity and specificity other than its advantages.Therefore,it’s of great significance to find a tumor biomarker with high sensitivity and specificity for the diagnosis of HCC.Researchers found out that LGALS3BP on exosome surface can be used as a biomarker for HCC.In this study,a detection system based on surface-enhanced Raman scattering(SERS)technology is designed to directly detect exosomal LGALS3BP in serum.In this system,the immunomagnetic beads(IMBs)capture exosomes by binding themselves to the marker protein CD63,and then the compounds are isolated under the action of magnetic field.In addition,the gold nanocages(AuNCs)Raman probe,the modification of anti-LG ALS 3 BP antibody(AuNCs-4MBA-anti-LGALS3BP),is formed into a compound that includes IMBs,exosomes and AuNCs Raman probes after the combination with LGALS3BP.The compound could generate SERS signals,and exosomes could be quantified according to SERS signals intensity.This subject mainly carried on the following researches.Firstly,we constructed the SERS biosensor consisting of capture probe based on IMBs and the detection probe based on AuNCs-4MBA-anti-LGALS3BP.And then we characterized them respectively.The results showed that the magnetic beads(MBs)and AuNCs had a hollow cage structure,uniform particle size and regular morphology,and AuNCs-4MBA-anti-LGALS3BP probes had the strong SERS activity.Secondly,we extracted exosomes secreted by HepG2 cells(HepG2-exos)and HL-7702 cells(HL-7702-exos)from cell culture supernatant,and conducted a test with the already-built SERS biosensor to investigate its sensitivity and specificity for the detection of HepG2-exos.The experimental results showed that,there was a good linear relationship between SERS intensity and the logarithmic value of HepG2-exos concentration when the HepG2-exos concentration was from 5×103 to 5×106 particles/mL.Besides,the SERS biosensor detected a significantly higher intensity of SERS signal generated by HepG2-exos than HL-7702-exos in the same concentration which indicated the strong specificity of the system for detecting HCC-derived exosomes.Thirdly,we detected exosomes in human serum samples.We measured serum samples from the healthy control group and the HCC patient group by our SERS biosensor,and evaluated the diagnostic efficacy of using exosomal LGALS3BP as the HCC biomarker.The results suggested a statistically significant difference between the two sets of measured data.The area under the receiver operating characteristic(AUROC)curves of the measured results was 0.94,indicating the high accuracy of exosomal LGALS3BP as the biomarker in HCC diagnosis.The experimental method of this study requires only 20μL serum samples to diagnose HCC,and the determination results could be obtained in 2 h.So it has great potential for a more sensitive and convenient diagnosis of HCC.