Yen-Ling Chen

Relying on a single biomarker in biomedical analysis is often insufficient for accurate disease or pathogen determination. A recent trend is using simultaneous multiplex detection of multiple biomarkers to improve diagnostic accuracy and throughput. To enable multiplex detection, we developed a series of surface-enhanced Raman scattering (SERS) nanoprobes, referred to as nanoaggregate-embedded beads (NAEBs). These NAEBs were synthesized using three distinct Raman reporter molecules: Safranin O, ethyl violet, and cresyl violet acetate. By integrating the NAEBs with magnetic nanoparticles and a simple capillary magnetofluidic device, we developed a rapid and simultaneous multiplex detection platform for genetic analysis of an aquacultural pathogen Vibrio parahaemolyticus (VP) for pirA, pirB, and ompA and genotyping of Alzheimer’s disease’s risk factor biomarker Apoliproprotein E (ApoE). For VP detection, a limit of detection (LOD) as low as ~ 10⁻¹⁶ M and a broad dynamic range of six orders were found . For ApoE genotyping, the platform was capable of detecting single nucleotide polymorphisms (SNPs) of rs429358 and rs7412 in the ApoE gene with a LOD of approximately 10⁻¹³ M. These results highlight the potential of this SERS-based simultaneous multiplex detection platform for sensitive and quantitative determination of multiple DNA biomarkers as well as SNPs in complex biological matrices. Graphical abstract

A fluorescent aptasensor was developed based on target-induced hairpin conformation switch coupled with nicking enzyme-assisted signal amplification (NESA) to detect the oligomeric form of ß-amyolid peptide (AβO) in cerebrospinal fluid. The hairpin DNA probe (HP) was specifically designed to recognize AβO. When AβO is present in the sensing system, it induces an HP conformational switch and triggers the NESA reaction. After evaluating the parameters of the aptasensor system, we selected Hairpin10, which has a 10-nucleotide extended sequence, as the hairpin sequence that interacts with AβO. The quantitative linear range of the proposed aptasensor is from 11.3 to 113 ng mL⁻¹ in artificial cerebrospinal fluid (aCSF), and the detection limit was 7.29 ng mL⁻¹. The present work realized the assay of AβO in aCSF with satisfactory quantitative results. Graphical abstract

Digoxin, a cardiac glycoside drug, is commonly used to treat heart failure and arrhythmias. The therapeutic concentration range of digoxin, with a narrow therapeutic index, is between 0.5 and 2.0 ng mL⁻¹. Hence, it is important for patients to monitor their blood levels after taking medication to achieve effective treatment and reduce the likelihood of experiencing drug side effects. Due to the complex steps and high cost of immunoassays, aptasensors that use aptamers to recognize the targets offer the advantages of low cost and good stability over other analysis methods. Nicking enzyme-assisted signal amplification is a novel isothermal signal amplification technology that relies on nicking enzymes to recognize and cleave restriction sites on one oligonucleotide strand. In this study, we develop a fluorescent aptasensor coupled with target-triggered aptamer hairpin switch and nicking enzyme-assisted signal amplification for digoxin detection in plasma for therapeutic drug monitoring. After optimizing the experimental parameters, we design hairpin probes with ten base pairs of the aptamer sequence and extended sequence complement to react with digoxin in a 10 mM Tris buffer containing 150 mM NaCl and 50 mM MgCl₂ (pH 7.4). The signal amplification reactions were performed for 3 hours. The fluorescent aptasensor exhibited high sensitivity with a detection limit of 88 pg mL⁻¹ for detecting digoxin in plasma and a linear range from 0.1 ng mL⁻¹ to 5 ng mL⁻¹. This technology was successfully used for digoxin detection to improve treatment effectiveness and minimize the risk of adverse side effects.

Alzheimer's disease (AD) is a chronic and progressive neurodegenerative disorder that affects millions of individuals worldwide. Researchers have conducted numerous studies to find accurate biomarkers for early AD diagnosis and develop more effective treatments. The main pathological hallmarks of AD are amyloid beta and Tau proteins. Other biomarkers, such as DNA, RNA, and proteins, can also be helpful in early AD diagnosis. To diagnose and treat AD promptly, it is essential to accurately measure the concentration of biomarkers in the cerebrospinal fluid or blood. However, due to the low concentrations of these biomarkers in the body, highly sensitive analytical techniques are required. To date, sensors have become increasingly important due to their high sensitivity, swift detection, and adaptable manipulation features. These qualities make them an excellent substitute for conventional instruments. Nanomaterials are commonly employed in sensors to amplify signals and improve sensitivity. This review paper summarized the integration of nanomaterials in optical sensor systems, including colorimetric, fluorescent, and surface-enhanced Raman scattering sensors for AD biomarkers detection.

Alzheimer's disease is a progressive neurodegenerative condition that causes brain cell death and is the leading cause of dementia. Most patients with Alzheimer's disease are diagnosed with late-onset Alzheimer's disease (LOAD), with apolipoprotein E (APOE) genotypes being highly associated with the frequency of LOAD risk. A fluorescence detection system coupled with oligonucleotide ligation and magnetic separation was developed to identify two single-nucleotide polymorphisms (SNPs) for the APOE gene and recognize APOE alleles for LOAD. The system utilized a fluorescence probe with one base-discriminating nucleoside for SNP (F probe) and a perfectly complementary biotin-modified sequence against the target DNA (P probe). When the F and P probes matched the target DNA sequences, DNA ligation occurred, and ligation products were produced. Streptavidin magnetic beads were subsequently employed to remove the ligation products, and a decrease in fluorescence intensity was observed in the supernatant compared to when there was no target DNA. This system detected two SNPs of APOE alleles, namely rs429358 and rs7412. The results indicated that the R-values ((F₀ − F₁)/F₀) for rs429358 were 0.92 ± 0.002 for the T/T target, 0.47 ± 0.004 for the T/C target and 0.11 ± 0.004 for the C/C target, respectively. The R-values for rs7412 were 0.73 ± 0.009 for the C/C target, 0.42 ± 0.001 for the C/T target and 0.16 ± 0.007 for the T/T target, respectively. F₀ and F₁ represent the fluorescence intensity of the F probe without and with target DNA, respectively. Based on fluorescence intensity, the fluorescence detection system was able to identify the genotypes of the APOE gene accurately to evaluate the risk of Alzheimer's disease.

Aβ oligomers (AβO) are a dominant biomarker for early Alzheimer’s disease diagnosis. A fluorescent aptasensor coupled with conformational switch–induced hybridization was established to detect AβO. The fluorescent aptasensor is based on the interaction of fluorophore-labeled AβO-specific aptamer (FAM-Apt) against its partly complementary DNA sequence on the surface of magnetic beads (cDNA-MBs). Once the FAM-Apt binds to AβO, the conformational switch of FAM-Apt increases the tendency to be captured by cDNA-MBs. This causes a descending fluorescence of supernatant, which can be utilized to determine the levels of AβO. Thus, the base-pair matching above 12 between FAM-Apt and cDNA-MBs with increasing hybridizing free energies reached the ascending fluorescent signal equilibrium. The optimized aptasensor showed linearity from 1.7 ng mL⁻¹ to 85.1 (R = 0.9977) with good recoveries (79.27–109.17%) in plasma. Furthermore, the established aptasensor possesses rational selectivity in the presence of monomeric Aβ, fibrotic Aβ, and interferences. Therefore, the developed aptasensor is capable of quantifying AβO in human plasma and possesses the potential to apply in clinical cases. Graphical Abstract