Vancomycin-modified Glass Slides Were Used For The Detection Of Staphylococcus Aureus By Surface-enhanced Raman Spectroscopy

TargetStaphylococcus aureus(S.aureus)is one of the common pathogenic bacteria,which can lead to skin and soft tissue infection,sepsis and catheter-related infection.As a light scattering technology,Surface-Enhanced Raman spectroscopy(SERS)has great potential in the field of clinical microbiological detection because of its advantages such as fast,real-time,non-destructive,easy to operate and less water interference.Vancomycin(Van)is a glycopeptide antibiotic.Its amino group forms a hydrogen bond complex with D-Ala-D-Ala at the acyl end of peptidoglycan promoter of bacterial cell wall to capture bacteria.It can be used as a biological recognition platform for bacterial capture.In this study,vancomycin was used as a carrier to capture S.aureus,and a detection method of S.aureus based on SERS was established.Method(1)The preparation of vancomycin modified slide:The surface of microscope cover glass was hydroxylated by 25%Na OH,and the hydroxylated glass were incubated in anhydrous ethanol of 1%3-aminopropyltriethoxysilane(APTES)for 12h to make the slide aminated,and then treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride(EDC),Vancomycin modified slide was prepared by covalent combination of carboxyl group of vancomycin and amino group of slide.Fourier transform infrared spectroscopy(FTIR)was used to characterize whether vancomycin was successfully modified.(2)Determination of vancomycin on glass slide:A certain concentration of vancomycin solution was set up.The absorbance of the vancomycin was determined by UV spectrophotometer and the standard curve of vancomycin concentration and UV absorbance was established.The UV absorbance changes of vancomycin solution before and after modification were measured,and the content of vancomycin on the surface of the slide was calculated according to the standard curve of vancomycin concentration and UV absorbance value(3)Capture of Staphylococcus aureus:The Staphylococcus aureus solution was dripped onto the surface of vancomycin modified slide and blank slide,incubated in 37?for 1 h,washed,dried,then observed by scanning electron microscope.(4)Determination of Staphylococcus aureus by SERS:By laser micro Raman spectroscopy at 633 nm,the SERS spectra of blank vancomycin modified slide,Staphylococcus aureus without silver nanoparticles and Staphylococcus aureus dropping with silver nanoparticles were detected.Result(1)Vancomycin modified slide:Compared with blank slide and aminated slide,vancomycin modified slide has infrared characteristic peak of benzene ring at 1538cm^-1,while aminated slide has amino characteristic peak at 1614 cm^-1,and blank slide has no obvious characteristic peak.(2)Vancomycin content on glass slide:The standard curve of vancomycin concentration and UV absorbance was established,Y=2.257X+0.126(Y refers to UV absorbance,X refers to vancomycin concentration).The vancomycin content on the surface of the modified slide was 0.02mg/cm².(3)Capture of Staphylococcus aureus on slide:Under the scanning electron microscope,it was observed that Staphylococcus aureus was successfully captured on the vancomycin modified slide,but not on the blank slide.(4)SERS spectra of Staphylococcus aureus:Under the laser micro Raman spectrometer,Staphylococcus aureus has SERS characteristic peaks at 724cm^-1,780cm^-1,1001cm^-1,1050 cm^-1,1100 cm^-1,1240 cm^-1.ConclusionStaphylococcus aureus was successfully captured and immobilized by vancomycin modified slide.With the enhancement of silver nanoparticles,the detection of Staphylococcus aureus was realized by SERS,which provided a basis for the direct identification of Staphylococcus aureus in human body fluids.