Label-free breast cancer detection and classification by convolutional neural network-based on exosomes surface-enhanced raman scattering

Because the breast cancer is an important factor that threatens women's lives and health,early diagnosis is helpful for disease screening and a good prognosis.Exosomes are nanovesicles,secreted from cells and other body fluids,which can reflect the genetic and phenotypic status of parental cells.Compared with other methods for early diagnosis of cancer(such as circulating tumor cells(CTCs)and circulating tumor DNA),exosomes have a richer number and stronger biological stability,and have great potential in early diagnosis.Thus,it has been proposed as promising biomarkers for diagnosis of early-stage cancer.However,distinguishing different exosomes remain is a major biomedical challenge.In this paper,we used predictive Convolutional Neural model to detect and analyze exosomes of normal and cancer cells with surface-enhanced Raman scattering(SERS).As a result,it can be seen from the SERS spectra that the exosomes of MCF-7,MDA-MB-231 and MCF-10A cells have similar peaks(939,1145 and 1380 cm^(-1)).Based on this dataset,the predictive model can achieve 95%accuracy.Compared with principal component analysis(PCA),the trained CNN can classify exosomes from different breast cancer cells with a superior performance.The results indicate that using the sensitivity of Raman detection and exosomes stable presence in the incubation period of cancer cells,SERS detection combined with CNN screening may be used for the early diagnosis of breast cancer in the future.

Application of microfluidic technology based on surface-enhanced Raman scattering in cancer biomarker detection:A review

With the continuous discovery and research of predictive cancer-related biomarkers,liquid biopsy shows great potential in cancer diagnosis.Surface-enhanced Raman scattering(SERS)and microfluidic technology have received much attention among the various cancer biomarker detection methods.The former has ultrahigh detection sensitivity and can provide a unique fingerprint.In contrast,the latter has the characteristics of miniaturization and integration,which can realize accurate control of the detection samples and high-throughput detection through design.Both have the potential for point-of-care testing(POCT),and their combination(lab-on-a-chip SERS(LoC-SERS))shows good compatibility.In this paper,the basic situation of circulating proteins,circulating tumor cells,exosomes,circulating tumor DNA(ctDNA),and microRNA(miRNA)in the diagnosis of various cancers is reviewed,and the detection research of these biomarkers by the LoC-SERS platform in recent years is described in detail.At the same time,the challenges and future development of the platform are discussed at the end of the review.Summarizing the current technology is expected to provide a reference for scholars engaged in related work and interested in this field.

Surface-enhanced Raman Scattering of Aflatoxin B1 on Silver by DFT Method

The structure, electrostatic properties, and Raman spectra of aflatoxin B1 （AFB1） and AFB1-Ag complex are studied by density functional theory with B3LYP/6- 311G（d,p）/Lan12dz basis set. The results show that the surface-enhanced Raman scattering （SERS） and pre-resonance Raman spectra of AFB1-Ag complex strongly depend on the adsorption site and the excitation wavelength found to enhance 102-103 order compared to of the incident light. The SERS factors are normal Raman spectrum of AFB1 molecule due to the larger static polarizabilities of the AFB1-Ag complex, which directly results in the stronger chemical enhancement in SERS spectra. The pre-resonance Raman spectra of AFB1-Ag complex are explored at 266, 482, 785, and 1064 nm incident light wavelength, in which the enhancement factors are about 10^2-10^4, mainly caused by the charge-transfer excitation resonance. The vibrational modes are analyzed to explain the relationship between the vibrational direction and the enhanced Raman intensities.

Surface-enhanced vibrational spectroscopies in electrocatalysis:Fundamentals,challenges,and perspectives

Electrocatalysis offers a promising approach towards chemical synthesis driven by renewable energy.Molecular level understanding of the electrochemical interface remains challenging due to its compositional and structural complexity.In situ interfacial specific characterization techniques could help uncover structure-function relationships and reaction mechanism.To this end,electrochemical surface-enhanced Raman spectroscopy(SERS)and surface-enhanced infrared absorption spectroscopy(SEIRAS)thrive as powerful techniques to provide fingerprint information of interfacial species at reaction conditions.In this review,we first introduce the fundamentals of SERS and SEIRAS,followed by discussion regarding the technical challenges and potential solutions.Finally,we highlight future directions for further development of surface-enhanced spectroscopic techniques for electrocatalytic studies.

Chemical Enhancement on Surface-Enhanced Resonance Raman Scattering of Au3-1,4-BenzenedithioI-Au3 Junction

Surface-enhanced Raman scattering （SERS） and surface-enhanced resonance Raman scattering （SERRS） spectra of the 1,4-benzenedithiol molecule in the junction of two Au3 clusters have been calculated using density functional theory （DFT） and time-dependent DFT method. In order to investigate the contribution of charge transfer （CT） enhancement, the wavelengths of incident light are chosen to be at resonance with four representative excited states, which correspond to CT in four different forms. Compared with SERS spectrum, SERRS spectra are enhanced enormously with distinct enhancement factors, which can be attributed to CT resonance in different forms.

Optical penetration of surface-enhanced micro-scale spatial offset Raman spectroscopy in turbid gel and biological tissue

The limited penetration of photons in biological tissue restricts the deep-tissue detection and imaging application.The micro-scale spatially offset Raman spectroscopy(micro-SORS)with an optical fiber probe,colleting photons from deeper regions by offsetting the position of laser excitation from the collection optics in a range of hundreds of microns,shows great potential to be integrated with endoscopy for inside-body noninvasive detection by circumventing this restric-tion,particularly with the combination of surface-enhanced Raman spectroscopy(SERS).However,a detailed tissue penetration study of micro-SORS in combination with SERS is still lacking.Herein,we compared the signal decay of enhanced Raman nanotags through the tissue phantom of agarose gel and the biological tissue of porcine muscle in the near-infrared(NIR)region using a portable Raman spectrometer with a micro-SORS probe(2.1 mm in diameter)and a conventional hand-held probe(9.7mm in diameter).Two kinds of Raman nanotags were prepared from gold nanorods decorated with the nonresonant(4-nitrobenzenethiol)or resonant Raman reporter molecules(IR-780 iodide).The SERS measurements show that the penetration depths of two Raman nanotags are both over 2 cm in agarose gel and 3 mm in porcine muscle.The depth could be improved to over 4 cm in agarose gel and 5 mm in porcine tissue when using the micro-SORS system.This demonstrates the superiority of optical-fiber micro-SORS system over the conventional Raman detection for the detection of nanotags in deeper layers in the turbid medium and biological tissue,offering the possibility of combining the micro-SORS technique with SERS for noninvasive in vivo endoscopy-integrated clinical application.

The isolation and identification of apolipoprotein C-I in hormone-refractory prostate cancer using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry

Androgens play a central role in prostate cancer pathogenesis, and hence most of the patients respond to androgen deprivation therapies. However, patients tend to relapse with aggressive prostate cancer, which has been termed as hormone refractory. To identify the proteins that mediate progression to the hormone-refractory state, we used protein-chip technology for mass profiling of patients＇ sera. This study included 16 patients with metastatic hormone-refractory prostate cancer who were initially treated with androgen deprivation therapy. Serum samples were collected from each patient at five time points： point A, pre-treatment; point B, at the nadir of the prostate- specific antigen （PSA） level; point C, PSA failure; point D, the early hormone-refractory phase; and point E, the late hormone-refractory phase. Using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry, we performed protein mass profiling of the patients＇ sera and identified a 6 640-Da peak that increased with disease progression. Target proteins were partially purified, and by amino acid sequencing the peak was identified as a fragment of apolipoprotein C-I （ApoC-I）. Serum ApoC-I protein levels increased with disease progression. On immunohistochemical analysis, the ApoC-i protein was found localized to the cytoplasm of the hormone-refractory cancer cells. In this study, we showed an increase in serum ApoC-I protein levels in prostate cancer patients during their progression to the hormone-refractory state, which suggests that ApoC-I protein is related to progression of prostate cancer. However, as the exact role of ApoC-I in prostate cancer pathogenesis is unclear, further research is required.

Surface-enhanced Raman spectroscopy of serum predicts sensitivity to docetaxel-based chemotherapy in patients with metastatic castration-resistant prostate cancer

Docetaxel-based chemotherapy,as the first-line treatment for metastatic castration-resistant prostate cancer(mCRPC),has succeeded in helping quite a number of patients to improve quality of life and prolong survival time.However,almost half of mCRPC patients are not sensitive to docetaxel chemotherapy initially.This study aimed to establish models to predict sensitivity to docetaxel chemotherapy in patients with mCRPC by using serum surface-enhanced Raman spectroscopy(SERS).A total of 32 mCPRC patients who underwent docetaxel chemo-therapy at our center from July 2016 to March 2018 were included in this study.Patients were dichotomized in prostate-specific antigen(PSA)response group(n=17)versus PSA failure group(n=15)according to the response to docetaxel.In total 64 matched spectra from 32 mCRPC patients were obtained by using SERS of serum at baseline(q0)and after 1 cycle of docetaxel chemotherapy(ql).Comparing Raman peaks of serum samples at baseline(q0)be-tween two groups,significant differences revealed at the peaks of 638,810,890(p<0.05)and 1136cm^(-1)(p<0.01).The prediction models of peak 1363 cm^(-1)and principal component anal-ysis and linear discriminant analysis(PCA-LDA)based on Raman data were established,re-spectively.The sensitivity and specificity of the prediction models were 71%,80%and 69%,78%through the way of leave-one-out cross-validation.According to the results of five-cross-valida-tion,the PCA-LDA model revealed an accuracy of 0.73 and AUC of 0.83.

Quantitative and sensitive detection of prohibited fish drugs by surface-enhanced Raman scattering

Rapid and simple detections of two kinds of prohibited fish drugs, crystal violet （CV） and malachite green （MG）, were accomplished by surface-enhanced Raman scattering （SERS）. Based on the optimized Au/cicada wing, the detectable concentration of CV/MG can reach 10-7 M, and the linear logarithmic quantitative relationship curves between log/and logC allows for the determination of the unknown concentration of CV/MG solution. The detection of these two analytes in real environment was also achieved, demonstrating the application potential of SERS in the fast screening of the prohibited fish drugs, which is of great benefit for food safety and environmental monitoring.

Sensitive and Label-Free Detection of Protein Secondary Structure by Amide Ⅲ Spectral Signals using Surface-Enhanced Raman Spectroscopy

Proteins and peptides perform a vital role in living systems, however it remains a challenge for accurate description of proteins at the molecular level. Despite that surface-enhanced Raman spectroscopy (SERS) can provide the intrinsic fingerprint information of samples with ultrahigh sensitivity, it suffers from the poor reproducibility and reliability. Herein, we demonstrate that the silver nanorod array fabricated by an oblique angle deposition method is a powerful substrate for SERS to probe the protein secondary structures without exogenous labels. With this method, the SERS signals of two typical proteins (lysozyme and cytochrome c) are successfully obtained. Additionally, by analyzing the spectral signals of the amide Ⅲ of protein backbone, the influence of concentration on the folding status of proteins has been elucidated. With the concentration increasing, the components of α-helix and β-sheet structures of lysozyme increase while the secondary structures of cytochrome c almost keep constant. The SERS method in this work offers an effective optical marker to characterize the structures of proteins.

Two-dimensional gel electrophoresis and surface-enhanced laser desorption ionization-time of flight-mass spectrometry for detection of protein expression profiles in the hippocampus following closed brain injury

Gene expression profile changes in brain regions following traumatic brain injury at the gene level cannot sufficiently elucidate gene expression time, expression amount, protein post-translational processing or modification. Therefore, it is necessary to quantitatively analyze the gene expression profile using proteomic techniques. In the present study, we established a rat model of closed brain injury using Marmarou＇s weight-drop device, and investigated hippocampal differential protein expression using two-dimensional gel electrophoresis and surface-enhanced laser desorption ionization-time of flight-mass spectrometry. A total of 364 protein peaks were detected on weak cation exchange-2 protein chips, including 37 differential protein peaks. 345 protein peaks were detected on immobilized metal affinity capture arrays-Cu, including 12 differential protein peaks Further examination of these differential proteins revealed that glucose-regulated protein and proteasome subunit alpha type 3 expression were significantly upregulated post-injury. These results indicate that brain injury can alter protein expression in the hippocampus, and that glucose-regulated protein and proteasome subunit alpha type 3 are closely associated with the occurrence and development of traumatic brain injury.

Surface-enhanced resonance Raman scattering spectroscopy of single R6G molecules

Surface-enhanced resonance Raman scattering （SERRS） of Rhodamine 6G （R6G） adsorbed on colloidal silver clusters has been studied. Based on the great enhancement of the Raman signal and the quench of the fluorescence, the SERRS spectra of R6G were recorded for the samples of dye colloidal solution with different concentrations. Spectral inhomogeneity behaviours from single molecules in the dried sample films were observed with complementary evidences, such as spectral polarization, spectral diffusion, intensity fluctuation of vibrational lines and even ＂breathing＂ of the molecules. Sequential spectra observed from a liquid sample with an average of 0.3 dye molecules in the probed volume exhibited the expected Poisson distribution for actually measuring 0, 1 or 2 molecules. Difference between the SERRS spectra of R6G excited by linearly and circularly polarized light were experimentally measured.

Identification and analysis of serum samples by surface-enhanced Raman spectroscopy combined with characteristic ratio method and PCA for gastric cancer detection

This study aimed to explore the application of surface-enhanced Raman scattering(SERS)in the rapid diagnosis of gastric cancer.The SERS spectra of 68 serum samples from gastric cancer patients and healthy volunteers were acquired.The characteristic ratio method(CRM)and principal component analysis(PCA)were used to differentiate gastric cancer serum from normal serum.Compared with healthy volunteers,the serum SERS intensity of gastric cancer patients was relatively high at 722 cm^(-1),while it was relatively low at 588,644,861,1008,1235,1397,1445 and 1586 cm^(-1).These results indicated that the relative content of nucleic acids in the serum of gastric cancer patients rises while the relative content of amino acids and carbohydrates decreases.In PCA,the sensitivity and specificity of discriminating gastric cancer were 94.1%and 94.1%,respectively,with the accuracy of 94.1%.Based on the intensity ratios of four characteristic peaks at 722,861,1008 and 1397 cm^(-1),CRM presented the diagnostic sensitivity and specificity of 100%and 97.4%,respectively,and the accuracy of 98.5%.Therefore,the three peak intensity ratios of I_(722)/I_(861),I_(722)/I_(1008)and I_(722)/I_(1397)can be considered as biologicalfingerprint information for gastric cancer diagnosis and can rapidly and directly reflect the physiological and pathological changes associated with gastric cancer development.This study provides an important basis and standards for the early diagnosis of gastric cancer.

Controllable Silver Plating on Silica for Surface-enhanced Raman Scattering

We proposed a facile and rapid method for preparing silica-silver core-shell（SSCS） substrates to use Ag electroless plating on SiO2@Au-seed particles.UV-Vis-NIR absorption spectrometer and SEM were employed to monitor the reaction process of the formation of Ag on the surfaces of silica beads,and the optical resonance of the substrate could shift from visible to NIR region.It has been found that surface-enhanced Raman scattering（SERS） enhancement changes with the electroless plating time and the SSCS substrate with the plating time of 90 s（90SSCS） shows the strongest SERS response under the laser excitation at 514.5 nm.Signals collected over multiple spots and substrate of rhodamine 6G（R6G） resulted in a relative standard deviation（RSD） of 9.75%.The calculated enhancement factor（EF） was approximately 105 ＂106.SSCS substrate exhibits high SERS performance,which is due to electromagnetic SERS enhancement with additional localization field within closely packed Ag nanoparticles decorated on the SiO2 nanoparticles.And this substrate presents tunable and broad localized surface plasmon resonance（LSPR）,so this method may open a new way for SERS studies with other laser excitation.

Molybdenum Nanoscrews:A Novel Non-coinage-Metal Substrate for Surface-Enhanced Raman Scattering

For the first time, Mo nanoscrew was cultivated as a novel non-coinage-metal substrate for surface-enhanced Raman scattering(SERS). It was found that the nanoscrew is composed of many small screw threads stacking along its length direction with small separations. Under external light excitation, strong electromagnetic coupling was initiated within the gaps, and many hot-spots formed on the surface of the nanoscrew, which was confirmed by high-resolution scanning near-field optical microscope measurements and numerical simulations using finite element method. These hotspots are responsible for the observed SERS activity of the nanoscrews. Raman mapping characterizations further revealed the excellent reproducibility of the SERS activity. Our findings may pave the way for design of low-cost and stable SERS substrates.

Raman and Surface-enhanced Raman Scattering of Chlorophenols

Raman spectrum is a powerful analytical tool for determining the chemical information of compounds. In this study, we obtained analytical results of chlorophenols（CPs） molecules including 4-chlorophenol（4-CP）, 2,6-dich- lorophenol（2,6-DCP） and 2,4,6-trichlorophenol（2,4,6-TCP） on the surface of Ag dendrites by surface-enhanced Raman scattering（SERS） spectra. SEM images indicate that the SERS substrate of Ag dendrites is composed of a large number of polygonal nanocrystallites, which self-assembled into a 3D hierarchical structure. It was found that there were distinct differences for those three molecules from Raman and SERS spectra. This indicates that SERS could be a new tool of detection technique regarding trace amounts of CPs.

Surface-enhanced Raman scattering on nanodiamond-derived carbon onions

Annealing nanodiamonds(ND) at high temperatures up to 1700 ℃ is a common method to synthesize carbon onions. The transformation from NDs to carbon onions is particularly interesting because of carbon onions' potential in the field of tribology and their application in ultra-charge/discharge devices. In this paper, a novel surface-enhanced Raman scattering technique that involves coating the sample with nanoscopic gold particles is proposed to characterize the NDs after different annealing treatments. Conventional Raman and surfaceenhanced Raman spectra were obtained, and the changes of peak parameters as the function of annealing temperature were evaluated. It was found that the widths of the D and the G peaks decreased with increasing annealing temperature, reflecting an improved order in the sp^2-hybridized carbon during the transformation from NDs to carbon onions. After annealing at 1700 ℃, the sp^2?carbon was highly ordered, indicating desirable electrical conductivity and lubricity. With increasing annealing temperature, the D peak showed a blue shift of almost30 cm^(-1), while the G peak merely shifted by 5 cm^(-1). For annealing temperatures above 1100 ℃, an increase of intensity ratio ID/IGwas observed. Compared to the uncoated area, red shifts of 0.5-2 cm^(-1) and of 5-9 cm^(-1) for the G and D peaks, respectively, were detected for the gold-coated area, which was due to the coupling of the plasmons and the phonons of the samples.

A new semi-quantitative Surface-Enhanced Raman Spectroscopy (SERS) method for detection of maleimide (2,5-pyrroledione) with potential application to astrobiology

Nitrogen-containing heterocyclic compounds are fundamental biochemical components of all life on Earth and,presumably,life elsewhere in our solar system.Detection and characterization of these compounds by traditional solvent extraction,chromatographic separation,and GC-MS analysis require more sample mass than will be available from samples returned to Earth from Mars.With its small sample mass requirement,Surface Enhanced Raman Spectroscopy could be an appropriate technique for analysis of returned samples.We have developed a SERS method for the detection of maleimide(2,5-pyrroledione),an N-containing heterocycle with a structure that is widespread in biochemicals.This semi-quantitative methodology accurately determines maleimide concentration in the range from 60 mg/mL to 120 mg/mL.We present a maleimide SERS standard spectrum which will be useful as a reference for future works.The present work demonstrates an easy,accurate,and effective method for the non-destructive qualitative and semi-quantitative study of maleimide as a first step toward developing a method for analysis of related compounds.

Highly Effective Detection of Amitraz in Honey by Using Surface-Enhanced Raman Scattering Spectroscopy Coupled with Chemometric Methods

As an effective and universal acaricide, amitraz is widely used on beehives against varroasis caused by the mite Varroa jacobsoni. Its residues in honey pose a great danger to human health. In this study, a sensitive, rapid, and environmentally friendly surface-enhanced Raman spectroscopy method (SERS) was developed for the determination of trace amount of amitraz in honey with the use of silver nanorod (AgNR) array substrate. The AgNR array substrate fabricated by an oblique angle deposition technique exhibited an excellent SERS activity with an enhancement factor of -10^7. Density function theory was employed to assign the characteristic peak of amitraz. The detection of amitraz was further explored and amitraz in honey at concentrations as low as 0.08 mg/kg can be identified. Specifically, partial least square regression analysis was employed to correlate the SERS spectra in full-wavelength with Camitraz to afford a multiple-quantitative amitraz predicting model. Preliminary results show that the predicted concentrations of amitraz in honey samples are in good agreement with their real concentrations. Compared with the conventional univariate quantitative model based on single peak’s intensity, the proposed multiple-quantitative predicting model integrates all the characteristic peaks of amitraz, thus offering an improved detecting accuracy and anti-interference ability.

Preparation of nano-structured Ag solid materials and application to surface-enhanced Raman scattering

Silver nano-particles with average diameter of about 60 nm were compacted in a high-strength mold under different pressures at 523 K to produce nano-structured Ag solid materials. The structure and characteristic of the nano-structured Ag solid materials (NSS-Ag) were studied using X-ray diffraction (XRD), scanning electron microscope (SEM) and Raman spectrometer. The NSS-Ag could be used as highly efficient surface-enhanced Raman scattering (SERS) active substrates. The common probe molecules Rhodamine 6G (R6G, 1×10-10 mol/L) were used to test the SERS activity on these substrates at very low concentrations. It is found that the SERS enhancement ability is dependent on the density of NSS-Ag. When the relative density of NSS-Ag is 83.87%, the materials reveal great SERS signal.