Search Results (11)

A bioluminescence-based assay for ATP can measure cell viability. Higher ATP concentration indicates a higher number of living cells. Thus, it is necessary to design an ATP sensor that is low-cost and easy to use. Gold nanoparticles provide excellent biocompatibility for enzyme immobilization. We investigated the effect of luciferase proximity with citrate-coated gold, silver, and gold–silver core–shell nanoparticles, gold nanorods, and BSA–Au nanoclusters. The effect of metal nanoparticles on the activity of luciferases was recorded by the luminescence assay, which was 3–5 times higher than free enzyme. The results showed that the signal stability in presence of nanoparticles improved and was reliable up to 6 h for analytes measurements. It has been suggested that energy is mutually transferred from luciferase bioluminescence spectra to metal nanoparticle surface plasmons. In addition, we herein report the 27-base DNA aptamer for adenosine-5′-triphosphate (ATP) as a suitable probe for the ATP biosensor based on firefly luciferase activity and AuNPs. Due to ATP application in the firefly luciferase reaction, the increase in luciferase activity and improved detection limits may indicate more stability or accessibility of ATP in the presence of nanoparticles. The bioluminescence intensity increased with the ATP concentration up to 600 µM with a detection limit of 5 µM for ATP. Full article

In this work, a label-free fluorescent detection method for glyphosate, based on DNA-templated silver nanoclusters (DNA-Ag NCs) and a Cu²⁺-ion-modulated strategy, was developed. In the presence of Cu²⁺, the fluorescence of the DNA-Ag NCs was quenched. Glyphosate can restore the fluorescence of DNA-Ag NCs. By analyzing the storage stability of the obtained DNA-Ag NCs using different DNA templates, specific DNA-Ag NCs were selected for the construction of the glyphosate sensor. The ultrasensitive detection of glyphosate was achieved by optimizing the buffer pH and Cu²⁺ concentration. The sensing of glyphosate demonstrated a linear response in the range of 1.0–50 ng/mL. The limit of detection (LOD) was 0.2 ng/mL. The proposed method was successfully applied in the detection of glyphosate in a real sample, indicating its high application potential for glyphosate detection. Full article

Salmonella Typhimurium (S. Typhimurium) is a globally distributed foodborne pathogen, which can lead to outbreaks of foodborne infectious diseases. It is essential to guarantee food safety by timely and correct detection of S. Typhimurium. In this investigation, an original fluorescence aptasensor was constructed to detect S. Typhimurium rapidly and sensitively. Through the coupling of magnetic beads, aptamer, and gold nanoparticles (AuNPs), a fluorescence quenching system with a “sandwich structure” was established. The aptamer acted as a link, and its specific binding to S. Typhimurium could release AuNPs from the system. Meanwhile, fluorescent DNA-stabilized silver nanoclusters (DNA-AgNCs) were synthesized. The fluorescence intensity changes caused by the fluorescence resonance energy transfer between DNA-AgNCs and AuNPs were utilized to detect S. Typhimurium. The purposed aptasensor exhibited high selectivity and sensitivity with a linear response to S. Typhimurium, ranging from 3.7 × 10² to 3.7 × 10⁵ cfu/mL. The limit of detection (LOD) was estimated to be 98 cfu/mL within 2 h 10 min. In addition, this method showed excellent application for detection of S. Typhimurium in artificially contaminated milk, with LOD reaching 3.4 × 10² cfu/mL. Therefore, the developed fluorescence aptasensor has great potential to identify S. Typhimurium in foodstuffs. Full article

In this study, we examined properties of silver nanoclusters, which are AgNCs stabilized by DNA oligonucleotide scaffold containing G-quadruplex-forming sequences: human telomeric (Tel22) or thrombin-binding aptamer (TBA). Thus, we obtained two fluorescent probes abbreviated as Tel22C12-AgNCs and TBAC12-AgNCs, which were characterized using absorption, circular dichroism and fluorescence spectroscopy. Both probes emit green and red fluorescence. The presence of silver nanoclusters did not destabilize the formed G-quadruplexes. The structural changes of probes upon binding K⁺ or Na⁺ ions cause quenching in their red emission. Green emission was slightly quenched only in the case of Tel22C12-AgNCs; on the contrary, for TBAC12-AgNC’s green emission, we observed an increasing fluorescence signal. Moreover, the Tel22C12-AgNCs system shows not only a higher binding preference for K⁺ over Na⁺, but it was able to monitor small changes in K⁺ concentrations in the buffer mimicking extracellular conditions (high content of Na⁺ ions). These results suggest that Tel22C12-AgNCs exhibit the potential to monitor transmembrane potassium transport. Full article

Atomically precise silver nanoclusters (AgNCs) are small nanostructures consisting of only a few atoms of silver. The combination of AgNCs with cytosine-rich single-stranded oligonucleotides results in DNA-templated silver nanoclusters (DNA-AgNCs). DNA-AgNCs are highly luminescent and can be engineered with reproducible and unique fluorescent properties. Furthermore, using nucleic acids as templates for the synthesis of AgNCs provides additional practical benefits by expanding optical activity beyond the visible spectral range and creating the possibility for color tunability. In this study, we explore DNA oligonucleotides designed to fold into hairpin-loop (HL) structures which modulate optical properties of AgNCs based on the size of the loop containing different number of cytosines (HL-C_N). Depending on the size of the loop, AgNCs can be manufactured to have either single or multiple emissive states. Such hairpin-loop structures provide an additional stability for AgNCs and further control over the base composition of the loop, allowing for the rational design of AgNCs’ optical properties. We demonstrate the potential of AgNCs in detecting Hg²⁺ by utilizing the HL-C₁₃ design and its variants HL-T₂C₁₁, HL-T₄C₉, and HL-T₆C₇. The replacement of cytosines with thymines in the loop was intended to serve as an additional sink for mercury ions extending the detectable range of Hg²⁺. While AgNC@HL-T₀C₁₃ exhibits an interpretable quenching curve, AgNC@HL-T₆C₇ provides the largest detectable range of Hg²⁺. The results presented herein suggest that it is possible to use a rational design of DNA-AgNCs based on the composition of loop sequence in HL structures for creating biosensors to detect heavy metals, particularly Hg²⁺. Full article

Silver has a long history of antibacterial effectiveness. The combination of atomically precise metal nanoclusters with the field of nucleic acid nanotechnology has given rise to DNA-templated silver nanoclusters (DNA-AgNCs) which can be engineered with reproducible and unique fluorescent properties and antibacterial activity. Furthermore, cytosine-rich single-stranded DNA oligonucleotides designed to fold into hairpin structures improve the stability of AgNCs and additionally modulate their antibacterial properties and the quality of observed fluorescent signals. In this work, we characterize the sequence-specific fluorescence and composition of four representative DNA-AgNCs, compare their corresponding antibacterial effectiveness at different pH, and assess cytotoxicity to several mammalian cell lines. Full article

Silver nanoclusters (AgNCs) generated on DNA templates belong to a new class of fluorescent tags showing excellent brightness, photostability and biocompatibility. Thus, AgNCs-DNA has been applied in various applications, from the detection of DNA/RNA and environmental monitoring to bioimaging and cancer therapy. In this work, we report fluorescent AgNCs synthesized using two 1,3-diaza-2-oxophenothiazine (tC)-modified oligonucleotides that contain RET-related sequence CCCCGCCCCGCCCCGCCCCA. The communication compares the absorption and emission properties of the obtained systems with silver nanoclusters synthesized on the unmodified oligonucleotide. First, we showed the optimal conditions for AgNCs-DNA synthesis on three DNA templates: (1) RET20 with the sequence 5′-CCC CGC CCC GCC CCG CCC CA-3′; (2) RET19tC with the sequence 5′-CCC CGC CCC GCC CCG CCC tCA-3′; and (3) RET14tC with the sequence 5′-CCC CGC CCC GCC CtCG CCC CA-3′. Next, the silver nanoclusters were characterized by UV/Vis absorption, fluorescence and circular dichroism spectroscopy. Silver nanoclusters RET19tC-AgNCs and RET14tC-AgNCs indicated several times higher fluorescence intensities in the long-wave emission spectra as compared to RET20-AgNCs. Moreover, silver nanoclusters on tC-modified oligonucleotides showed higher stability over time. The possibility of using the silver nanoclusters RET19tC-AgNCs for monitoring pH changes will be also tested. Full article

DNA-encapsulated Silver Nanoclusters (DNA/AgNCs) based sensors have gained increasing attention in past years due to their diverse applications in bioimaging, biosensing, and enzymatic assays. Given the potential of DNA/AgNCs for practical applications, the systematic studies of the fluorescent stability over an extended period is necessary. However, the correlation between nucleic acid properties and the long-term stability of DNA/AgNCs is less known. With locking-to-unlocking sensors, in which the secondary structure of DNA template is standardized, we investigated the correlation between the DNA structure and the fluorescence stability of AgNCs. Post-synthesis of DNA/AgNCs, the fluorescence, and structures of templates were monitored over three weeks. By combining the fluorescence spectroscopy with the in-gel fluorescent assay, we found that AgNCs encapsulated by dimer-structured DNA/AgNCs templates were more stable than those of hairpin-structured DNA/AgNCs templates. While the orange fluorescence from the dimer templates increased over three weeks, the red fluorescence from the hairpin templates was diminished by >80% within two days at room temperature. Further tests revealed that hairpin-encapsulated red-emissive AgNCs is more sensitive to oxidation by atmospheric oxygen compared to dimer encapsulated orange AgNCs. Our observations may provide an important clue in encapsulating photophysically more stable AgNCs by tuning the DNA secondary structures. The proposed strategy here can be essential for pragmatic applications of DNA/AgNCs templates. Full article

Nucleic acid stability and structure, which are crucial to the properties of fluorescent DNA-templated silver nanoclusters (DNA-Ag NCs), significantly change in ionic liquids. In this work, our purpose was to study DNA-Ag NCs in a buffer containing the hydrated ionic liquid of choline dihydrogen phosphate (choline dhp) to improve fluorescence for application in DNA detection. Due to the stabilisation of an i-motif structure by the choline cation, a unique fluorescence emission—that was not seen in an aqueous buffer—was observed in choline dhp and remained stable for more than 30 days. A DNA-Ag NCs probe was designed to have greater fluorescence intensity in choline dhp in the presence of a target DNA. A turn-on sensing platform in choline dhp was built for the detection of the BRCA1 gene, which is related to familial breast and ovarian cancers. This platform showed better sensitivity and selectivity in distinguishing a target sequence from a mutant sequence in choline dhp than in the aqueous buffer. Our study provides new evidence regarding the effects of structure on properties of fluorescent DNA-Ag NCs and expands the applications of fluorescent DNA-Ag NCs in an ionic liquid because of improved sensitivity and selectivity. Full article

Trypsin is important during the regulation of pancreatic exocrine function. The detection of trypsin activity is currently limited because of the need for the substrate to be labeled with a fluorescent tag. A label-free fluorescent method has been developed to monitor trypsin activity. The designed peptide probe consists of six arginine molecules and a cysteine terminus and can be conjugated to DNA-stabilized silver nanoclusters (DNA-AgNCs) by Ag-S bonding to enhance fluorescence. The peptide probe can also be adsorbed to the surface of graphene oxide (GO), thus resulting in the fluorescence quenching of DNA-AgNCs-peptide conjugate because of Förster resonance energy transfer. Once trypsin had degraded the peptide probe into amino acid residues, the DNA-AgNCs were released from the surface of GO, and the enhanced fluorescence of DNA-AgNCs was restored. Trypsin can be determined with a linear range of 0.0–50.0 ng/mL with a concentration as low as 1 ng/mL. This label-free method is simple and sensitive and has been successfully used for the determination of trypsin in serum. The method can also be modified to detect other proteases. Full article

Unmodified single-stranded DNA has recently gained popularity for the templated synthesis of fluorescent noble metal nanoclusters (NCs). Bright, stable, and biocompatible clusters have been developed primarily through optimization of DNA sequence. However, DNA backbone modifications have not yet been investigated. In this work, phosphorothioate (PS) DNAs are evaluated in the synthesis of Au and Ag nanoclusters, and are employed to successfully template a novel emitter using T₁₅ DNA at neutral pH. Mechanistic studies indicate a distinct UV-dependent formation mechanism that does not occur through the previously reported thymine N3. The positions of PS substitution have been optimized. This is the first reported use of a T₁₅ template at physiological pH for AgNCs. Full article