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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. [ABSTRACT FROM AUTHOR]

In this work, Au nanobowl (AuNB) arrays and the hybridization chain reaction (HCR) were utilized to develop a surface-enhanced Raman scattering (SERS) microfluidic chip for the ultrasensitive and simultaneous detection of squamous cell carcinoma antigen (SCCA) and cytokeratin 19 fragment antigen 21–1 (CYFRA21–1) in the serum of cervical cancer patients. Antibody-oligonucleotide conjugates were adopted as triggers, which initiated HCR by the hybridization between hairpin DNAs that were modified on Raman reporters-labeled Ag nanocubes, forming nicked double-stranded DNAs on AuNB arrays and leading to SERS signal amplification. Good linear response in the range of 10-12 – 10-6 g/mL, with an ultralow limit of detection for SCCA (0.08 pg/mL) and CYFRA21–1 (0.13 pg/mL), was achieved. Furthermore, the experiments demonstrated that the chip was feasible, time-efficient, and selective. The proposed chip was successfully applied to detect SCCA and CYFRA21–1 in clinical samples, and the results were consistent with those of enzyme-linked immunosorbent assay. Since most protein biomarkers in serum lack the corresponding aptamers, this method, which establishes antibody-oligonucleotide conjugates to trigger HCR, provides a general signal amplification technology for detecting protein biomarkers and exhibits good potential in clinical applications. • A SERS microfluidic chip based on AuNBs array and HCR was developed and employed for clinical detection. • Antibody-oligonucleotide conjugates were established to connect with targets and initiate HCR for signal amplification. • The assay possessed high sensitivity, with LODs as low as 0.08 pg/mL for SCCA and 0.13 pg/mL for CYFRA21-1. • The SERS microfluidic chip realized automatic flow by hydrophilic treatment and comb-like structure channels. [ABSTRACT FROM AUTHOR]

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. • CHA-induced powerful cycle amplification and the aggregation effect caused by MBs significantly enhanced the sensitivity of proposed pump-free SERS microfluidic chip, with the LODs low to aM level. • The pump-free SERS microfluidic chip presented satisfactory uniformity, stability, specificity and reproducibility. • This dual-signal amplification strategy could be applied for the dynamic monitoring expression of ctDNA in mice serum at different stages. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Non-small cell lung cancer (NSCLC) is a major histologic category of lung cancer. In this paper, a new type of surface-enhanced Raman scattering (SERS) microfluidic chip proposed for the simultaneous and efficient detection of miR-92a and miR-339-3p, two microRNAs (miRNAs) related to NSCLC. In addition, the method was used to detect miR-92a and miR-339-3p in the serum of NSCLC clinical samples, and the results were consistent with the quantitative real-time polymerase chain reaction (qRT-PCR). In conclusion, this new SERS microfluidic chip is a very promising tool for detecting miRNA expression levels, which can help in the early diagnosis and prognosis of NSCLC. [Display omitted] • SERS effect of the Au-AgNBs array is excellent, which improves the sensitivity of SERS analysis. • The hydrophilic-treated SERS microfluidic chip combined with the hpDNA- functioned Au-AgNBs array eliminates the need for an external driver while improving detection efficiency. • The LOD of the SERS microfluidic chip reaches aM level, and the detection and analysis time is as low as 8 min. • The device can be used for the early diagnosis of non-small cell lung cancer to improve cancer survival rates. In this paper, a novel surface-enhanced Raman scattering (SERS) microfluidic chip is proposed for the simultaneous and efficient detection of miR-92a and miR-339-3p, two microRNAs (miRNAs) related to non-small cell lung cancer (NSCLC). The sample solution can flow along the microchannel into the reaction zone during the assay, driven by the capillary force generated by the comb-like structured channel. A one-step paired hybridization reaction with the corresponding hairpin DNA (hpDNA) on the hpDNA-functioned Au-Ag nanobowl (Au-AgNBs) array takes place when the target strand is present in the sample solution, amplifying the SERS signal. The detection limits of miR-92a and miR-339-3p in serum were 44.36 aM and 63.58 aM, respectively, using this method, with a detection and analysis time of only 8 min. The microfluidic chip showed good specificity, reproducibility, and stability. Finally, the method was used to detect miR-92a and miR-339-3p in the serum of NSCLC clinical samples, and the results were consistent with the quantitative real-time polymerase chain reaction (qRT-PCR). In conclusion, this new SERS microfluidic chip is a very promising tool for detecting miRNA expression levels, which can help in the early diagnosis and prognosis of NSCLC. [ABSTRACT FROM AUTHOR]

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 pump-free SERS microfluidic chip coupling with CHA amplification is fabricated for the ultrasensitive and simultaneous detection of miR-106b and miR-196b. [Display omitted] • To our knowledge, this is the first report demonstrating pump-free SERS microfluidic chip coupling with catalytic hairpin assembly (CHA) reaction for detection of miR-106b and miR-196b simultaneously. • Dual signal amplification composed of CHA-induced powerful cycle amplification and the aggregation effect caused by MBs enables the ultrasensitive detection of miRNA, with the LODs low to aM level. • The entire detection process could be finished less than 5 min. • Excellent uniformity, stability, specificity and reproducibility of this pump-free SERS microfluidic chip are demonstrated. [ABSTRACT FROM AUTHOR]