A SERS Biosensor Based on Functionalized Au-SiNCA Integrated with a Dual Signal Amplification Strategy for Sensitive Detection of Telomerase Activity During EMT in Laryngeal Carcinoma.

Purpose: This paper aims to construct a surface-enhanced Raman spectroscopy (SERS) biosensor based on functionalized Au-Si nanocone arrays (Au-SiNCA) using a dual signal amplification strategy (SDA-CHA) to evaluate telomerase activity during epithelial-mesenchymal transition (EMT) in laryngeal carcinoma (LC). Methods: A SERS biosensor based on functionalized Au-SiNCA was designed with an integrated dual-signal amplification strategy to achieve ultrasensitive detection of telomerase activity during EMT in LC patients. Results: Labeled probes (Au-AgNRs@4-MBA@H1) and capture substrates (Au-SiNCA@H2) were prepared by modifying hairpin DNA and Raman signal molecules. Using this scheme, telomerase activity in peripheral mononuclear cells (PMNC) could be successfully detected with a limit of detection (LOD) as low as 10-6 IU/mL. In addition, biological experiments using BLM treatment of TU686 effectively mimicked the EMT process. The results of this scheme were highly consistent with the ELISA scheme, confirming its accuracy. Conclusion: This scheme provides a reproducible, selective, and ultrasensitive assay for telomerase activity, which is expected to be a potential tool for the early screening of LC in future clinical applications.

Association between follicle-stimulating hormone and nonalcoholic fatty liver disease in postmenopausal women with type 2 diabetes mellitus.

BACKGROUND AND AIM: Follicle-stimulating hormone (FSH) was negatively associated with nonalcoholic fatty liver disease (NAFLD) in women older than 55 years old. People with obesity and diabetes had higher prevalence of NAFLD. Thus, we aimed to explore the association between FSH and NAFLD in postmenopausal women with type 2 diabetes mellitus (T2DM). METHODS: A total of 583 postmenopausal women with T2DM with an average age of 60.22 ± 6.49 were recruited in this cross-sectional study through January 2017 to May 2021. Anthropological data, biochemical indexes, and abdominal ultrasound results were retrospectively collected. Abdominal ultrasound was used to diagnose NAFLD. FSH was measured by enzymatic immunochemiluminescence and divided into tertiles for further analysis. The logistic regression was used to assess the association of FSH with prevalent NAFLD. Likelihood ratio tests were used to assess the interactions between groups. RESULTS: A total of 332 (56.94%) postmenopausal women had NAFLD. Compared with postmenopausal women in the lowest tertile of FSH, postmenopausal women in the highest tertile of FSH had lower prevalence of NAFLD (p < .01). After adjusting for age, diabetes duration, metabolism-related indicators, and other sex-related hormones, FSH was inversely associated with NAFLD (odds ratio: 0.411, 95% confidence intervals: 0.260-0.651, p < .001). In subgroup analysis, there were no significant interactions of FSH with strata of metabolic factors on the association of NAFLD. CONCLUSION: FSH was negatively and independently associated with NAFLD in postmenopausal women with type 2 diabetes mellitus. It might be a potential index for screening and identifying individuals with high risk of NAFLD in postmenopausal women.

Au/SiNCA-based SERS analysis coupled with machine learning for the early-stage diagnosis of cisplatin-induced liver injury.

Cisplatin has been widely applied in the clinical treatment of various cancers, whereas liver injury induced by its hepatotoxicity is still a severe issue. Reliable identification of early-stage cisplatin-induced liver injury (CILI) can improve clinical care and help to streamline drug development. Traditional methods, however, cannot achieve enough information at the subcellular level due to the requirement of the labeling process and low sensitivity. To overcome these, we designed an Au-coated Si nanocone array (Au/SiNCA) to fabricate the microporous chip as the surface-enhanced Raman scattering (SERS) analysis platform for the early diagnosis of CILI. A CILI rat model was established, and the exosome spectra were obtained. The principal component analysis (PCA)-representation coefficient-based k-nearest centroid neighbor (RCKNCN) classification algorithm was proposed as the multivariate analysis method to build the diagnosis and staging model. The PCA-RCKNCN model has been validated to achieve a satisfactory result, with accuracy and AUC of over 97.5%, and sensitivity and specificity of over 95%, indicating that SERS combined with the PCA-RCKNCN analysis platform can be a promising tool for clinical applications.

LoC-SERS Platform Integrated with the Signal Amplification Strategy toward Parkinson's Disease Diagnosis.

Multiplexed detection of Parkinson's disease (PD) biomarkers is of great significance for early diagnosis and personalized treatment. In this study, we fabricated a robust surface-enhanced Raman scattering-enabled lab-on-a-chip (LoC-SERS) platform for simultaneous quantification of α-synuclein, phosphorylated tau protein 181, osteopontin, and osteocalcin. Herein, the antibody-DNA conjugate was designed to introduce the catalytic hairpin self-assembly (CHA) amplification into the protein detection. Au nano-stars (AuNSs) modified with Raman reporter molecules and hairpin-structure DNA 1 were applied as the SERS nanotags. Au-coated silicon nanocone array (Au/SiNCA) fabricated based on the maskless plasma etching-prepared high-density Si nanocone array (SiNCA) and surface ion sputtering was used as the capture substrate after the modification of hairpin-structure DNA 2. Benefitting from the antibody-DNA conjugate-induced CHA amplification, numerous AuNSs can be connected to the Au/SiNCA surface, which significantly amplify the plasmonic coupling effect for ultrasensitive SERS detection, and the limit of detection was less than the pg/mL level. The application of highly uniform Au/SiNCA and antibody-DNA conjugate endows the LoC-SERS platform excellent analytical performance, including superior reproducibility, satisfactory universality, and high sensitivity. In addition, a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice model was established, and satisfactory results were obtained in real sample analysis with the LoC-SERS platform, which may be enlightening for exploiting protein biomarkers in PD monitoring.

Multiplex signal amplification strategy-based early-stage diagnosis of Parkinson's disease on a SERS-enabled LoC system.

In this paper, a multiplex signal amplification strategy was developed for the determination of miR-214 and miR-221 on a surface-enhanced Raman scattering (SERS)-enabled lab-on-a-chip (LoC) system to realize the early-stage diagnosis of Parkinson's disease (PD). The gold nanobipyramids (GNBPs) with great monodispersity were functionalized with Raman reporter molecules and hairpin DNA 1, serving as the SERS nanotags. The presence of targets can initial the strand displacement amplification (SDA) reaction and the numerous short-stranded trigger DNA (tDNA) can be released under the action of polymerase and nicking enzyme. Then, the tDNA can trigger the catalytic hairpin assembly (CHA) event between the SERS nanotags and the capture nanoprobes (Magnetic beads (MBs) modified with hairpin DNA 2), resulting in the aggregation of GNBPs on the MBs surface. The multiplex signal amplification contributed by the SDA-CHA strategy and the magnet-induced aggregation effect can ultimately lead to the significant improvement of the detection sensitivity and the limit of detection (LOD) was low to aM level with reproducibility and specificity meanwhile. Furthermore, a MPTP-induced PD mice model was established to verify the practicability and the expression level of miR-214 and miR-221 at different stages analyzed with the LoC system was confirmed by qRT-PCR.

A capillary-driven LoC-SERS device integrated with catalytic hairpin assembly amplification technology for NSCLC-related biomarkers detection.

In this study, we apply catalytic hairpin assembly (CHA) as the signal amplification strategy for the quantification of carcinoembryonic antigen (CEA) and cytokeratin fragment antigen 21-1 (CYFRA21-1) with a surface-enhanced Raman scattering (SERS) microfluidic chip (LoC-SERS) as the carrier. Herein, antibody-DNA conjugates are designed to assist the application of CHA amplification in protein detection. In the presence of protein biomarkers, antibody-DNA conjugates can specifically bind to the target proteins, forming the antigen@antibody-DNA conjugates. The terminal free part of the DNA on the conjugates can trigger the CHA events to connect SERS nanotags to capture nanoprobes. Then, micro-magnet can gather the CHA products in a rectangular chamber, resulting in the aggregation of SERS nanotags, which can ultimately generate abundant "hot spots" for SERS signal enhancement. Using this strategy, CEA and CYFRA21-1 can be successfully determined with a limit of detection (LOD) as low as pg mL-1, much lower than recently reported methods. Meanwhile, a non-small cell lung cancer (NSCLC)-xenografted mouse model was established, and SERS was applied to analyze the expression level of CEA and CYFRA21-1 in tumorigenesis and development. The comparison between SERS results and those of the ELISA method demonstrated a high degree of consistency, suggesting that the proposed CHA-assisted LoC-SERS device has satisfying accuracy. Thus, introducing the CHA strategy via the design of antibody-DNA conjugates opens new gates to ultra-sensitive and specific SERS detection of protein biomarkers.

A microfluidic chip using Au@SiO2 array-based highly SERS-active substrates for ultrasensitive detection of dual cervical cancer-related biomarkers.

In this work, a microfluidic chip using Au@SiO2 array-based highly active SERS substrates was developed for quantitative detection of squamous cell carcinoma antigen (SCCA) and carcinoembryonic antigen (CEA) associated with cervical cancer. The chip consisted of six functional units with pump-free design, enabling parallel detection of multiple samples in an automatic manner without external pumps and improving the portability. Ag nanocubes (AgNCs) were labeled with Raman reporters and coupled with antibodies (labeling) to prepare SERS tags, while the Au nanoparticle-modified SiO2 microsphere (Au@SiO2) array was conjugated with antibodies (coating) to generate the highly SERS-active capturing substrate. In the presence of target biomarkers, they were captured by SERS tags and capturing substrate, resulting in the formation of "sandwich" structures which were trapped in the detection chamber. As the immune reaction proceeded, a large number of "hot spots" were generated by the proximity of the Au@SiO2 array substrate and AgNCs, greatly amplifying SERS signals. With this chip, the limits of detection of the SCCA and CEA levels in human serum were estimated to be as low as 0.45 pg mL-1 and 0.36 pg mL-1, respectively. Furthermore, the good selectivity and reproducibility of this chip were confirmed. Finally, clinical serum samples were analyzed by this chip, and the outcomes were consistent with those of enzyme-linked immunosorbent assay (ELISA). Thus, the proposed microfluidic chip can be potentially applied for the clinical diagnosis of cervical cancer.

Simultaneous detection of circulating tumor DNAs using a SERS-based lateral flow assay biosensor for point-of-care diagnostics of head and neck cancer.

Circulating tumor DNA (ctDNA) has recently emerged as an ideal target for biomarker analytes. Thus, the development of rapid and ultrasensitive ctDNA detection methods is essential. In this study, a high-throughput surface-enhanced Raman scattering (SERS)-based lateral flow assay (LFA) strip is proposed. The aim of this method is to achieve accurate quantification of TP53 and PIK3CA E545K, two types of ctDNAs associated with head and neck squamous cell carcinoma (HNSCC), particularly for point-of-care testing (POCT). Raman reporters and hairpin DNAs are used to functionalize the Pd-Au core-shell nanorods (Pd-AuNRs), which serve as the SERS probes. During the detection process, the existence of targets could open the hairpins on the surface of Pd-AuNRs and trigger the first step of catalytic hairpin assembly (CHA) amplification. The next stage of CHA amplification is initiated by the hairpins prefixed on the test lines, generating numerous "hot spots" to enhance the SERS signal significantly. By the combination of high-performing SERS probes and a target-specific signal amplification strategy, TP53 and PIK3CA E545K are directly quantified in the range of 100 aM-1 nM, with the respective limits of detection (LOD) calculated as 33.1 aM and 20.0 aM in the PBS buffer and 37.8 aM and 23.1 aM in human serum, which are significantly lower than for traditional colorimetric LFA methods. The entire detection process is completed within 45 min, and the multichannel design realizes the parallel detection of multiple groups of samples. Moreover, the analytical performance is validated, including reproducibility, uniformity, and specificity. Finally, the SERS-LFA biosensor is employed to analyze the expression levels of TP53 and PIK3CA E545K in the serum of patients with HNSCC. The results are verified as consistent with those of qRT-PCR. Thus, the SERS-LFA biosensor can be considered as a noninvasive liquid biopsy assay for clinical cancer diagnosis.

Highly sensitive and simultaneous detection of ctDNAs related to non-small cell lung cancer in serum using a catalytic hairpin assembly strategy in a SERS microfluidic chip.

Circulating tumor DNA (ctDNA) is an ideal biomarker for cancer diagnosis based on liquid biopsy, so there is an urgent need for developing an efficient, rapid, and ultrasensitive detection method to meet clinical needs. In this paper, a novel surface-enhanced Raman scattering (SERS) microfluidic chip combined with a catalytic hairpin assembly (CHA) was proposed to detect two non-small cell lung cancer (NSCLC)-related ctDNA (TP53 and PIK3CA-Q546K) simultaneously. The chip consists of six channels for parallel detection. In the reaction region, the CHA reaction between HP1 of the SERS probe and HP2 of the capture substrate was triggered by ctDNAs to form HP1-HP2 duplexes. As the reaction proceeds, more and more SERS probes are captured on the substrate. The gathered reaction products continuously form a lot of hot spots, which greatly enhance the SERS signal. This reaction was completed within 5 minutes. Through this method, the detection limits of TP53 and PIK3CA-Q546K in human serum were as low as 2.26 aM and 2.34 aM, respectively. The microfluidic chip also exhibited high specificity, reproducibility and stability. The clinical feasibility of the SERS microfluidic chip was verified by analyzing the serum samples of healthy subjects and NSCLC patients. The reliability of the experimental results was verified by the qRT-PCR test. The constructed SERS-based analytical micro-platform has great potential in dynamic monitoring of cancer staging and could be used as a clinical tool for early cancer screening.

A pump-free and high-throughput microfluidic chip for highly sensitive SERS assay of gastric cancer-related circulating tumor DNA via a cascade signal amplification strategy.

Circulating tumour DNA (ctDNA) has emerged as an ideal biomarker for the early diagnosis and prognosis of gastric cancer (GC). In this work, a pump-free, high-throughput microfluidic chip coupled with catalytic hairpin assembly (CHA) and hybridization chain reaction (HCR) as the signal cascade amplification strategy (CHA-HCR) was developed for surface-enhanced Raman scattering (SERS) assays of PIK3CA E542K and TP53 (two GC-related ctDNAs). The chip consisted of six parallel functional units, enabling the simultaneous analysis of multiple samples. The pump-free design and hydrophilic treatment with polyethylene glycol (PEG) realized the automatic flow of reaction solutions in microchannels, eliminating the dependence on external heavy-duty pumps and significantly improving portability. In the reaction region of the chip, products generated by target-triggered CHA initiated the HCR, forming long nicked double-stranded DNA (dsDNA) on the Au nanobowl (AuNB) array surface, to which numerous SERS probes (Raman reporters and hairpin DNA-modified Cu2O octahedra) were attached. This CHA-HCR strategy generated numerous active "hot spots" around the Cu2O octahedra and AuNB surface, significantly enhancing the SERS signal intensity. Using this chip, an ultralow limit of detection (LOD) for PIK3CA E542K (1.26 aM) and TP53 (2.04 aM) was achieved, and the whole process was completed within 13 min. Finally, a tumour-bearing mouse model was established, and ctDNA levels in mouse serum at different stages were determined. To verify the experimental accuracy, the gold-standard qRT-PCR assay was utilized, and the results showed a high degree of consistency. Thus, this rapid, sensitive and cost-effective SERS microfluidic chip has potential as an ideal detection platform for ctDNA monitoring.

Pump-free microfluidic chip based laryngeal squamous cell carcinoma-related microRNAs detection through the combination of surface-enhanced Raman scattering techniques and catalytic hairpin assembly amplification.

MicroRNA (miRNA), as one of the ideal target biomarker analytes, plays an essential role in biological processes; thus, the development of rapidly sensitive detection methods is imperative. Herein, we proposed a pump-free surface-enhanced Raman scatting (SERS) microfluidic chip for the rapid and ultrasensitive detection of miR-106b and miR-196b, laryngeal squamous cell carcinoma (LSCC)-related miRNAs. Ag-Au core-shell nanorods (Ag-AuNRs) were applied to prepare SERS tags by modifying Raman reporters and hairpin DNAs. The capture probes were synthesized by labeling hairpin DNAs onto the magnetic beads (MBs) surface. In the presence of targets, the catalytic hairpin assembly (CHA) reactions between SERS tags and capture probes could be triggered, causing the aggregation of Ag-AuNRs. The tiny magnets installed under the rectangular chamber could magnetically gather the CHA products, leading to the further aggregation of Ag-AuNRs. Thus, this strategy could achieve the double aggregation of Ag-AuNRs, resulting in the significant amplification of the SERS signal. The proposed strategy achieved simultaneous and sensitive detection of miR-106b and miR-196b, with limits of detection low to aM level. The whole detection process could be completed within 5 min. Moreover, this microfluidic chip exhibited excellent reproducibility, stability, and specificity. The high accuracy of this SERS microfluidic chip was proved by practical analysis in LSCC patients' serum. The results demonstrated that SERS could be a promising alternative clinical diagnosis tool and exhibited potential application for the dynamic monitoring of cancer staging.

A dual-signal amplification strategy based on pump-free SERS microfluidic chip for rapid and ultrasensitive detection of non-small cell lung cancer-related circulating tumour DNA in mice serum.

Circulating tumour DNAs (ctDNAs) have been reported to be associated with real-time information of progression; however, an accurate and sensitive method has not been established. Herein, a novel dual-signal amplification strategy based on a pump-free surface-enhanced Raman scattering (SERS) microfluidic chip and a catalytic hairpin assembly (CHA) technique was developed for the dynamic monitoring of BRAF V600E and KRAS G12V, which are two non-small cell lung cancer (NSCLC)-related ctDNAs. In the presence of targets, CHA reactions can be triggered between two hairpin DNAs, fixing Pd-Au core-shell nanorods (Pd-AuNRs) onto the magnetic beads (MBs) surface. Thereafter, the composite structures can assemble under the action of magnet, enabling dual-amplification of SERS signal. Using this strategy, the limit of detection (LOD) was low (i.e. at the aM level) in serum. Furthermore, the entire chip-based analysis process could be completed within 5 min, eliminating manual incubation and heavy-duty injection pumps. The selectivity, reproducibility and uniformity of the proposed pump-free SERS microfluidic chip were satisfactory. This superior analysis strategy was finally applied to quantify BRAF V600E and KRAS G12V in tumour-bearing nude mice serum, the results of which corresponded with those of real-time polymerase chain reaction. Overall, this study provides a promising alternative tool for the dynamic monitoring of ctDNA expression level which can benefit the clinical diagnosis of NSCLC.

A dual-amplification strategy-intergated SERS biosensor for ultrasensitive hepatocellular carcinoma-related telomerase activity detection.

Telomerase has been considered as a biomarker for early diagnosis and prognosis assessment of hepatocellular carcinoma (HCC), while the highly sensitive and specific methods remain challenging. To detect telomerase, a novel surface-enhanced Raman scattering (SERS) biosensor was constructed using the dual DNA-catalyzed amplification strategy composed of strand displacement amplification (SDA) and catalytic hairpin assembly (CHA). This strategy relies on the extension reaction of telomerase primer induced by telomerase, forming long-stranded DNAs with repetitive sequence to catalyze the follow-up SDA event. Subsequently, the SDA products can trigger the CHA reaction between the SERS probes (Au-Ag nanocages (Au-AgNCs) modified with hairpin DNA1 and Raman reporters) and capture substrate (Au@SiO2 array labeled with hairpin DNA2), resulting in the formation of numerous "hot spots" to significantly enhance the SERS signal. Results are promising that the established biosensor presented excellent reproducibility, specificity and sensitivity. Moreover, ELISA was applied as the golden standard to verify the application of the proposed biosensor in real samples and the results confirmed the satisfactory accuracy of our method. Therefore, the proposed SERS biosensor has the potential to be an ideal tool for the early screening of HCC.

A novel DNA biosensor for the ultrasensitive detection of DNA methyltransferase activity based on a high-density "hot spot" SERS substrate and rolling circle amplification strategy.

Herein, we proposed a novel biosensor based on a high-density "hot spot" Au@SiO2 array substrate and rolling circle amplification (RCA) strategy for the ultrasensitive detection of CpG methyltransferase (M.SssI) activity. In the presence of M.SssI, the RCA process can be triggered, causing the augmentation of the single-stranded DNA (ssDNA) at the tail of the double-stranded DNA (dsDNA), and the ssDNA can be hybridized with numerous DNA probes labeled with Raman reporters in the next steps. Afterwards, the resultant ssDNA can be modified to the Au@SiO2 array substrate with the SERS enhancement factor of 7.49 × 106. The substrate was synthesized by using a monolayer SiO2 array to pick up the Au nanoparticle (AuNP) array and finite-difference time-domain (FDTD) simulation showed its excellent SERS effect. Particularly, the developed biosensor displayed a significant sensitivity with a broad detection range covering from 0.005 to 50 U mL-1, and the limits of detection (LODs) in PBS buffer and human serum were 2.37 × 10-4 U mL-1 and 2.51 × 10-4 U mL-1, respectively. Finally, in order to verify the feasibility of its clinical application, the serum samples of healthy subjects and breast cancer, prostate cancer, gastric cancer and cervical cancer patients were analyzed, and the reliability of the results was also confirmed by western blot (WB) experiments. Taking advantage of these merits, the proposed biosensor can be a very promising alternative tool for the detection of M.SssI activity, which is of vital importance in the early detection and prevention of tumors.

SERS Based Lateral Flow Assay for Rapid and Ultrasensitive Quantification of Dual Laryngeal Squamous Cell Carcinoma-Related miRNA Biomarkers in Human Serum Using Pd-Au Core-Shell Nanorods and Catalytic Hairpin Assembly.

Non-invasive early diagnosis is of great significant in disease pathologic development and subsequent medical treatments, and microRNA (miRNA) detection has attracted critical attention in early cancer screening and diagnosis. However, it was still a challenge to report an accurate and sensitive method for the detection of miRNA during cancer development, especially in the presence of its analogs that produce intense background noise. Herein, we developed a surface-enhanced Raman scattering (SERS)-based lateral flow assay (LFA) biosensor, assisted with catalytic hairpin assembly (CHA) amplification strategy, for the dynamic monitoring of miR-106b and miR-196b, associated with laryngeal squamous cell carcinoma (LSCC). In the presence of target miRNAs, two hairpin DNAs could self-assemble into double-stranded DNA, exposing the biotin molecules modified on the surface of palladium (Pd)-gold (Au) core-shell nanorods (Pd-AuNRs). Then, the biotin molecules could be captured by the streptavidin (SA), which was fixed on the test lines (T1 line and T2 line) beforehand. The core-shell spatial structures and aggregation Pd-AuNRs generated abundant active "hot spots" on the T line, significantly amplifying the SERS signals. Using this strategy, the limits of detections were low to aM level, and the selectivity, reproducibility, and uniformity of the proposed SERS-LFA biosensor were satisfactory. Finally, this rapid analysis strategy was successfully applied to quantitatively detect the target miRNAs in clinical serum obtained from healthy subjects and patients with LSCC at different stages. The results were consistent with the quantitative real-time PCR (qRT-PCR). Thus, the CHA-assisted SERS-LFA biosensor would become a promising alternative tool for miRNAs detection, which showed a tremendous clinical application prospect in diagnosing LSCC.

A novel surface-enhanced Raman scattering probe based on Au nanoboxes for dynamic monitoring of caspase-3 during cervical cancer cell apoptosis.

The highly sensitive and reliable detection, imaging, and monitoring of changes of intracellular caspase-3 are critical for understanding the cell apoptosis and studying the progression of caspase-3-related cervical cancer. Herein, we present a novel surface-enhanced Raman scattering (SERS) probe for the detection of caspase-3 during cervical cancer cell apoptosis, composed of Au nanoboxes modified with Nile blue A as a Raman reporter and a caspase-3-specified peptide as a molecular cross-linker. In the presence of caspase-3, the substrate peptides can be cleaved and the changed surface charge of the Au nanoboxes results in the Au nanoboxes-NBA-peptide assembling to form aggregates and a great enhancement of SERS signal. The finite-difference time-domain simulation showed that hot spots mainly located in the nanogaps of the aggregated Au nanoboxes, which in theory proved the rationality of this signal amplification method. The SERS probes exhibited excellent reproducibility and selectivity toward caspase-3. A detection limit of 0.127 fM was obtained for caspase-3, with a dynamic range from 1 fM to 1 nM. MTT assay demonstrated that the probes had no obvious cytotoxicity within a certain concentration range. HeLa cells were treated with doxorubicin to induce long-term apoptosis. Upon cellular uptake of these probes, the spatiotemporal dynamics of caspase-3 in apoptotic cells could be real-time monitored using SERS. The activity of caspase-3 increased with the prolongation of apoptosis time. The SERS results were in accordance with that of western blotting assay. This kind of probe can offer great potential for the determination of enzymatic activities in the physiological processes of cells.

Highly sensitive detection of cytochrome c in the NSCLC serum using a hydrophobic paper based-gold nanourchin substrate.

Cytochrome c (Cyt c) is a biomarker of early apoptosis that plays a critical role in the diagnosis and therapy of non-small cell lung cancer (NSCLC). In this work, we proposed a novel surface-enhanced Raman scattering (SERS)-based biosensor to implement the ultrasensitive detection of Cyt c in the serum of NSCLC patients. The SERS-supporting substrates based on hydrophobic filter paper were composed of gold nanourchins (GNUs) surface-functionalized with the Cyt c aptamer and the cyanine 5-labeled complementary DNA. In the existence of Cyt c, it could specifically bind to its aptamer, which leads to the detachment of complementary strands modified with Cy5 and the great weakness of SERS signal. The finite-difference time domain (FDTD) simulation showed that the excellent SERS performance of GNUs aggregation was strongly dependent on a large number of "hot spots" at the tips and between the nanogaps of aggregated GNUs. Alkyl ketene dimer (AKD) was used to make the filter paper modify its property from hydrophilic to hydrophobic, which consequently increased the density of GNUs and extended the retention time of the analyte. SERS biosensors based on hydrophobic paper exhibited prominent reproducibility and selectivity. The detection limit of Cyt c in PBS was 1.148 pg/mL, while the detection limit in human serum was 1.79 pg/mL. Moreover, the analysis of the serum samples of healthy subjects and NSCLC patients confirmed the feasibility of its clinical application. The results were consistent with enzyme-linked immunosorbent assay results. This method can be a powerful strategy for quantitative detection of extracellular Cyt c, and it is expected that the SERS-based biosensors could be applied in the practical clinical diagnoses of NSCLC.

Role of Ketotifen on metabolic profiles, inflammation and oxidative stress in diabetic rats.

We aim to explore effects of Ketotifen on metabolic profiles, inflammation and oxidative stress. Sprague Dawley (SD) male rats were randomly divided into normal control group (NC) and experimental groups, and experimental group rats were fed with high-sugar and fat diet for 6 weeks. Then, experimental group rats were divided into diabetes group (DM) and ketotifen treatment group (KT). KT group was given ketotifen via Intragastric for 8 weeks with the dosage of 0.09 mg/kg/d. Fasting plasma glucose (FPG) was measured using glucose oxidase-phenol amino phenazone method. Fasting insulin (FINS), total cholesterol (TC), triglyceride (TG), low-density lipoprotein (LDL), high-density lipoprotein (HDL), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were tested by enzyme-linked immunosorbent assay. Malondialdehyde (MDA) and superoxide dismutase (SOD) were quantified by spectrophotometer method. Before Ketotifen administration, compared with NC group, DM and KT groups showed significantly high levels of body weight, FPG, FINS, HOMA-IR, TC, TG, LDL, IL-6, TNF-α and MDA, and lower levels of HDL and SOD (All p <0.05). After 4 weeks of Ketotifen administration, levels of body weight, FPG, FINS, HOMA-IR, TC, TG, LDL, IL-6, TNF-α in KT group decreased significantly, and levels of HDL and SOD elevated significantly (All p <0.05). After 8 weeks of Ketotifen administration, levels of body weight, FPG, FINS, HOMA-IR, TC, TG, LDL, IL-6, TNF-α and MDA in KT group decreased more obviously, and levels of HDL and SOD elevated significantly further (All p <0.05). Ketotifen improved metabolic profiles, and ameliorated status of inflammation and oxidative stress.

The Geriatric Nutritional Risk Index Independently Predicts Mortality in Diabetic Foot Ulcers Patients Undergoing Amputations.

Objective. Patients with diabetic foot ulcers undergoing amputations have poor prognosis. Malnutrition usually occurs in this population and is associated with increased risk of mortality. The geriatric nutritional risk index (GNRI) is a widely used, simple, and well-established tool to assess nutritional risk. The purpose of this study was to assess the association between GNRI and all-cause mortality in diabetic foot ulcers patients undergoing minor or major amputations. Methods. This was a retrospective cohort study including 271 adult patients. Patients were divided into two groups according to a GNRI cutoff value of 92, and characteristics and mortality were compared between the two groups. Cox proportional hazard analysis was performed to explore the association between GNRI and mortality. Result. GNRI (p < 0.001), age (p < 0.001), and eGFR (p = 0.002) were independent predictors of mortality. Among a subgroup of 230 patients with minor amputation, increased age (p < 0.001), coronary artery disease (p = 0.030), and increased GNRI (p < 0.001) were major risk factors. Conclusion. GNRI on admission might be a novel clinical predictor for the incidence of death in patients with diabetic foot ulcers who were undergoing amputations.

Associations of serum anti-ganglioside antibodies and inflammatory markers in diabetic peripheral neuropathy.

AIMS: To investigate the associations between inflammatory markers, serum anti-ganglioside antibodies (anti-GS-ab), serum plasminogen activator inhibitor-1 (PAI-1), tumor necrosis factor-α (TNF-α), C-reactive protein (CRP), and diabetic peripheral neuropathy (DPN). METHODS: Study subjects were divided into three groups: normal group (N group) with 101 healthy individuals; diabetes mellitus without peripheral neuropathy group (DM group) with 87 patients; and DPN group with 178 cases. American Nicolet Viking IV electromyography was applied to detect nerve conduction velocity; enzyme linked immunosorbent assay was used to determine the levels of anti-GS-IgG-ab, PAI-1, and TNF-α; and immunoturbidimetry was employed to measure CRP levels. RESULTS: Motor nerve conduction velocity and sensory nerve conduction velocity in the DNC group were significantly lower than in the N and DM groups (all P<0.05). Pairwise comparisons among diabetic peripheral neuropathy clinical (DPNC) levels were statistically significant (P<0.05), and the level of anti-GS-ab was positively correlated with DPNC. There were statistically significant differences in PAI-1, TNF-α, and CRP levels between the DPN group and DM and N groups (both P<0.05). Pairwise comparisons of PAI-1, TNF-α, and CRP levels among DPNC levels showed no statistical significance in volumes (P>0.05), and the concentration of anti-GS-IgM-ab was in significant positive correlated with PAI-1, TNF-α, and CRP levels. CONCLUSION: Anti-GS-ab and inflammatory markers such as PAI-1, TNF-α, and CRP were associated with DPN and can be used as important indicators for the prediction and early diagnosis of DPN.