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.

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.

No abstract

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.

Circulating tumor DNA (ctDNA) is an ideal biomarker for cancer diagnosis based on liquid biopsy, so there is an urgent need to develop an efficient, rapid, and ultrasensitive detection method...

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 conjugateinduced 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.

Herein, we proposed a novel biosensor based on the high-density “hot spots” Au@SiO2 array substrate and rolling circle amplification (RCA) strategy for ultrasensitive detection of the CpG methyltransferase (M.SssI) activity....

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.