Research

14 pages, 6450 KiB

Open AccessArticle

Broad-Spectrum Antibody-Based Immunochromatographic Strip Assay for Rapid Screening of Bisphenol A Diglycidyl Ether and Its Derivatives in Canned Foods

by Chundi Yu, Jinnuo Hu, Wei Wu, Yongfei Zhou, Can Zhang and Qingli Yang

Molecules 2024, 29(1), 13; https://doi.org/10.3390/molecules29010013 - 19 Dec 2023

Viewed by 630

Abstract

Bisphenol A diglycidyl ether (BADGE) is widely present in the inner coating of metal food cans, from which it can migrate into food and generate harmful derivatives during storage, such as bisphenol A (2,3-dihydroxypropyl) glycidyl ether, bisphenol A (3-chloro-2-hydroxypropyl) glycidyl ether, and bisphenol [...] Read more.

Bisphenol A diglycidyl ether (BADGE) is widely present in the inner coating of metal food cans, from which it can migrate into food and generate harmful derivatives during storage, such as bisphenol A (2,3-dihydroxypropyl) glycidyl ether, bisphenol A (3-chloro-2-hydroxypropyl) glycidyl ether, and bisphenol A (3-chloro-2-hydroxypropyl) (2,3-dihydroxypropyl) glycidyl ether. Here, a gold-nanoparticle-based immunochromatographic strip assay based on a broad-spectrum polyclonal antibody was developed for the simultaneous detection of BADGE and its derivatives, which could be accomplished within 15 min. The quantitative analysis of the visualization results was performed using Adobe Photoshop CC 2021, and the detection limit, defined as the concentration causing 15% inhibition, was 0.97 ng/mL. The recoveries of BADGE and its derivatives at various spiking levels in canned food samples ranged from 79.86% to 93.81%. The detection results of the proposed immunochromatographic strip assay were validated via high-performance liquid chromatography, showing a good correlation coefficient (R² = 0.9580). Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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19 pages, 2403 KiB

Open AccessCommunication

Systematic Analysis of 2′-O-Alkyl Modified Analogs for Enzymatic Synthesis and Their Oligonucleotide Properties

by Kenta Ishida, Yuuya Kasahara, Hidekazu Hoshino, Takumi Okuda and Satoshi Obika

Molecules 2023, 28(23), 7911; https://doi.org/10.3390/molecules28237911 - 02 Dec 2023

Viewed by 1105

Abstract

Enzymatic oligonucleotide synthesis is used for the development of functional oligonucleotides selected by in vitro selection. Expanding available sugar modifications for in vitro selection helps the functional oligonucleotides to be used as therapeutics reagents. We previously developed a KOD DNA polymerase mutant, KOD [...] Read more.

Enzymatic oligonucleotide synthesis is used for the development of functional oligonucleotides selected by in vitro selection. Expanding available sugar modifications for in vitro selection helps the functional oligonucleotides to be used as therapeutics reagents. We previously developed a KOD DNA polymerase mutant, KOD DGLNK, that enzymatically synthesized fully-LNA- or 2′-O-methyl-modified oligonucleotides. Here, we report a further expansion of the available 2′-O-alkyl-modified nucleotide for enzymatic synthesis by KOD DGLNK. We chemically synthesized five 2′-O-alkyl-5-methyluridine triphosphates and incorporated them into the oligonucleotides. We also enzymatically synthesized a 2′-O-alkyl-modified oligonucleotide with a random region (oligonucleotide libraries). The 2′-O-alkyl-modified oligonucleotide libraries showed high nuclease resistance and a wide range of hydrophobicity. Our synthesized 2′-O-alkyl-modified oligonucleotide libraries provide novel possibilities that can promote the development of functional molecules for therapeutic use. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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18 pages, 6159 KiB

Open AccessArticle

Electrochemiluminescence Aptasensor with Dual Signal Amplification by Silica Nanochannel-Based Confinement Effect on Nanocatalyst and Efficient Emitter Enrichment for Highly Sensitive Detection of C-Reactive Protein

by Ning Ma, Shuai Xu, Weidong Wu and Jiyang Liu

Molecules 2023, 28(22), 7664; https://doi.org/10.3390/molecules28227664 - 19 Nov 2023

Cited by 2 | Viewed by 980

Abstract

The rapid and sensitive detection of the important biomarker C-reactive protein (CRP) is of great significance for monitoring inflammation and tissue damage. In this work, an electrochemiluminescence (ECL) aptasensor was fabricated based on dual signal amplification for the sensitive detection of CRP in [...] Read more.

The rapid and sensitive detection of the important biomarker C-reactive protein (CRP) is of great significance for monitoring inflammation and tissue damage. In this work, an electrochemiluminescence (ECL) aptasensor was fabricated based on dual signal amplification for the sensitive detection of CRP in serum samples. The sensor was constructed by modifying a silica nanochannel array film (SNF) on a cost-effective indium tin oxide (ITO) electrode using the Stöber solution growth method. Gold nanoparticles (AuNPs) were grown in situ within the nanochannels using a simple electrodeposition method as a nanocatalyst to enhance the active electrode area as well as the ECL signal. The negatively charged nanochannels also significantly enriched the positively charged ECL emitters, further amplifying the signal. The recognition aptamer was covalently immobilized on the outer surface of SNF after modification with epoxy groups, constructing the aptasensor. In the presence of CRP, the formation of complexes on the recognitive interface led to a decrease in the diffusion of ECL emitters and co-reactants to the supporting electrode, resulting in a reduction in the ECL signal. Based on this mechanism, ECL detection of CRP was achieved with a linear range of 10 pg/mL to 1 μg/mL and a low limit of detection (7.4 pg/mL). The ECL aptasensor developed in this study offers advantages such as simple fabrication and high sensitivity, making promising applications in biomarker detection. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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21 pages, 2664 KiB

Open AccessArticle

Structure-Guided Development of Bivalent Aptamers Blocking SARS-CoV-2 Infection

by Md Shafiqur Rahman, Min Jung Han, Sang Won Kim, Seong Mu Kang, Bo Ri Kim, Heesun Kim, Chang Jun Lee, Jung Eun Noh, Hanseong Kim, Jie-Oh Lee and Sung Key Jang

Molecules 2023, 28(12), 4645; https://doi.org/10.3390/molecules28124645 - 08 Jun 2023

Cited by 3 | Viewed by 2041

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused devastation to human society through its high virulence, infectivity, and genomic mutations, which reduced the efficacy of vaccines. Here, we report the development of aptamers that effectively interfere with SARS-CoV-2 infection by targeting its [...] Read more.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused devastation to human society through its high virulence, infectivity, and genomic mutations, which reduced the efficacy of vaccines. Here, we report the development of aptamers that effectively interfere with SARS-CoV-2 infection by targeting its spike protein, which plays a pivotal role in host cell entry of the virus through interaction with the viral receptor angiotensin-converting enzyme 2 (ACE2). To develop highly effective aptamers and to understand their mechanism in inhibiting viral infection, we determined the three-dimensional (3D) structures of aptamer/receptor-binding domain (RBD) complexes using cryogenic electron microscopy (cryo-EM). Moreover, we developed bivalent aptamers targeting two distinct regions of the RBD in the spike protein that directly interact with ACE2. One aptamer interferes with the binding of ACE2 by blocking the ACE2-binding site in RBD, and the other aptamer allosterically inhibits ACE2 by binding to a distinct face of RBD. Using the 3D structures of aptamer–RBD complexes, we minimized and optimized these aptamers. By combining the optimized aptamers, we developed a bivalent aptamer that showed a stronger inhibitory effect on virus infection than the component aptamers. This study confirms that the structure-based aptamer-design approach has a high potential in developing antiviral drugs against SARS-CoV-2 and other viruses. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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11 pages, 2735 KiB

Open AccessArticle

Highly Sensitive β-Lactoglobulin Fluorescent Aptamer Biosensors Based on Tungsten Disulfide Nanosheets and DNase I-Assisted Signal Amplification

by Yuying Wang, Sisi Chen, Wanmei Chen, Jingjing Wang, Kun Li, Chengyi Hong, Kailong Zhang and Quansheng Chen

Molecules 2023, 28(8), 3502; https://doi.org/10.3390/molecules28083502 - 16 Apr 2023

Cited by 1 | Viewed by 1647

Abstract

β-lactoglobulin (β-Lg) is a protein found in milk that can cause severe allergic reactions, including rash, vomiting, and diarrhea. Thus, it is crucial to develop a sensitive β-Lg detection method to protect people who are susceptible to allergies. Here, we introduce a novel [...] Read more.

β-lactoglobulin (β-Lg) is a protein found in milk that can cause severe allergic reactions, including rash, vomiting, and diarrhea. Thus, it is crucial to develop a sensitive β-Lg detection method to protect people who are susceptible to allergies. Here, we introduce a novel and highly sensitive fluorescent aptamer biosensor for detecting β-Lg. First, a fluorescein-based dye (FAM)-labeled β-lactoglobulin aptamer (β-Lg aptamer) is adsorbed on the surface of tungsten disulfide (WS₂) nanosheets via van der Waals forces, resulting in fluorescence quenching. When β-Lg is present, the β-Lg aptamer selectively binds to β-Lg, causing a conformational change in the β-Lg aptamer and releasing it from the surface of WS₂ nanosheets, which restores the fluorescence signal. Simultaneously, DNase I in the system cleaves the aptamer bound to the target, producing a short oligonucleotide fragment and releasing β-Lg. The released β-Lg then binds to another β-Lg aptamer adsorbed on WS₂, initiating the next round of cleavage, resulting in significant amplification of the fluorescence signal. This method has a linear detection range of 1–100 ng mL⁻¹, and the limit of detection is 0.344 ng mL⁻¹. Furthermore, this approach has been successfully used for detecting β-Lg in milk samples with satisfactory results, providing new opportunities for food analysis and quality control. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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12 pages, 1843 KiB

Open AccessArticle

The SEB1741 Aptamer Is an Efficient Tool for Blocking CD4+ T Cell Activation Induced by Staphylococcal Enterotoxin B

by Leslie Chavez-Galan, Andy Ruiz, Lucero A. Ramón-Luing, Alejandro Escamilla-Gutiérrez, Anahí Sánchez-Monciváis, Brenda Tecuatzi-Cadena, Karen Medina-Quero and María Guadalupe Córdova-Espinoza

Molecules 2023, 28(8), 3480; https://doi.org/10.3390/molecules28083480 - 14 Apr 2023

Cited by 1 | Viewed by 1695

Abstract

Staphylococcal enterotoxin B (SEB) is a protein produced by Staphylococcus aureus, which is toxic to humans. It is well known for its ability to stimulate the exacerbated activation of proinflammatory CD4+ T cells (Th1 profile), and in vitro studies have been conducted [...] Read more.

Staphylococcal enterotoxin B (SEB) is a protein produced by Staphylococcus aureus, which is toxic to humans. It is well known for its ability to stimulate the exacerbated activation of proinflammatory CD4+ T cells (Th1 profile), and in vitro studies have been conducted to understand its mechanism of action and its potential use as an immune therapy. However, the efficiency of the SEB1741 aptamer in blocking SEB has not been experimentally demonstrated. Methods: Enrichment CD4+ T cells were stimulated with SEB, and as a blocker, we used the SEB1741 aptamer, which was previously synthesised by an “in silico” analysis, showing high affinity and specificity to SEB. The efficiency of the SEB1741 aptamer in blocking CD4+ T cell activation was compared with that of an anti-SEB monoclonal antibody. Flow cytometry and Bio-Plex were used to evaluate the T-cell function. Results: In vitro, SEB induced the activation of CD4+ T cells and favoured a Th1 profile; however, the SEB1741 aptamer was highly efficient in decreasing the frequency of CD4+ T cells positive to ki-67 and CD69 cells, this means that proliferation and activation of CD4+ T cells was decreased. Moreover, the production of interleukin 2 (IL-2) and interferon-gamma (IFN-γ) was affected, suggesting that the Th1 profile is not present when the SEB1441 aptamer is used. Thus, the SEB1741 function was similar to that of anti-SEB. Conclusions: The SEB1741 aptamer is a valuable tool for blocking CD4+ T cell activation and the subsequent release of proinflammatory cytokines by SEB stimulation. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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12 pages, 1473 KiB

Open AccessArticle

A Dual-Mode Method Based on Aptamer Recognition and Time-Resolved Fluorescence Resonance Energy Transfer for Histamine Detection in Fish

by Xin Wang, Fu Yang, Chengfang Deng, Yujie Zhang, Xiao Yang, Xianggui Chen, Yukun Huang, Hua Ye, Jianjun Zhong and Zhouping Wang

Molecules 2022, 27(24), 8711; https://doi.org/10.3390/molecules27248711 - 09 Dec 2022

Cited by 2 | Viewed by 1647

Abstract

Histamine produced via the secretion of histidine decarboxylase by the bacteria in fish muscles is a toxic biogenic amine and of significant concern in food hygiene, since a high intake can cause poisoning in humans. This study proposed a fluorometric and colorimetric dual-mode [...] Read more.

Histamine produced via the secretion of histidine decarboxylase by the bacteria in fish muscles is a toxic biogenic amine and of significant concern in food hygiene, since a high intake can cause poisoning in humans. This study proposed a fluorometric and colorimetric dual-mode specific method for the detection of histamine in fish, based on the fluorescence labeling of a histamine specific aptamer via the quenching and optical properties of gold nanoparticles (AuNPs). Due to the fluorescence resonance energy transfer phenomenon caused by the proximity of AuNPs and NaYF4:Ce/Tb, resulting in the quenching of the fluorescence signal in the detection system, the presence of histamine will compete with AuNPs to capture the aptamer and release it from the AuNP surface, inducing fluorescence recovery. Meanwhile, the combined detection of the two modes showed good linearity with histamine concentration, the linear detection range of the dual-mode synthesis was 0.2–1.0 μmol/L, with a detection limit of 4.57 nmol/L. Thus, this method has good selectivity and was successfully applied to the detection of histamine in fish foodstuffs with the recoveries of 83.39~102.027% and 82.19~105.94% for Trichiurus haumela and Thamnaconus septentrionalis, respectively. In addition, this method was shown to be simple, rapid, and easy to conduct. Through the mutual verification and combined use of the two modes, a highly sensitive, rapid, and accurate dual-mode detection method for the analysis of histamine content in food was established, thereby providing a reference for the monitoring of food freshness. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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15 pages, 2090 KiB

Open AccessArticle

Aptamer Selection Based on Microscale Electrophoretic Filtration Using a Hydrogel-Plugged Capillary Device

by Junku Takao, Reina Nagai, Tatsuro Endo, Hideaki Hisamoto and Kenji Sueyoshi

Molecules 2022, 27(18), 5818; https://doi.org/10.3390/molecules27185818 - 08 Sep 2022

Cited by 4 | Viewed by 2091

Abstract

This study reports a novel aptamer selection method based on microscale electrophoretic filtration. Aptamers are versatile materials that recognize specific targets and are attractive for their applications in biosensors, diagnosis, and therapy. However, their practical applications remain scarce due to issues with conventional [...] Read more.

This study reports a novel aptamer selection method based on microscale electrophoretic filtration. Aptamers are versatile materials that recognize specific targets and are attractive for their applications in biosensors, diagnosis, and therapy. However, their practical applications remain scarce due to issues with conventional selection methods, such as complicated operations, low-efficiency separation, and expensive apparatus. To overcome these drawbacks, a selection method based on microscale electrophoretic filtration using a capillary partially filled with hydrogel was developed. The electrophoretic filtration of model target proteins (immunoglobulin E (IgE)) using hydrogel, the electrokinetic injection of DNAs to interact with the trapped proteins, the elimination of DNAs with weak interactions, and the selective acquisition of aptamer candidates with strong interactions were successfully demonstrated, revealing the validity of the proposed concept. Two aptamer candidates for IgE were obtained after three selection cycles, and their affinity for the target was confirmed to be less than 1 nM based on their dissociation constant (K_D) values. Therefore, the proposed method allows for the selection of aptamers with simple operations, highly effective separation based on electrophoresis and filtration, and a relatively cheap apparatus with disposable devices. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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10 pages, 3054 KiB

Open AccessArticle

Evolution of Interferon-Gamma Aptamer with Good Affinity and Analytical Utility by a Rational In Silico Base Mutagenesis Post-SELEX Strategy

by Lianhui Zhao, Qionglin Wang, Yingai Yin, Yan Yang, Huifang Cui and Yiyang Dong

Molecules 2022, 27(17), 5725; https://doi.org/10.3390/molecules27175725 - 05 Sep 2022

Cited by 4 | Viewed by 2003

Abstract

The Systematic Evolution of Ligands by EXponential enrichment (SELEX) is conventionally an effective method to identify aptamers, which are oligonucleotide sequences with desired properties to recognize targets specifically and sensitively. However, there are some inherent limitations, e.g., the loss of potential high-affinity sequences [...] Read more.

The Systematic Evolution of Ligands by EXponential enrichment (SELEX) is conventionally an effective method to identify aptamers, which are oligonucleotide sequences with desired properties to recognize targets specifically and sensitively. However, there are some inherent limitations, e.g., the loss of potential high-affinity sequences during biased iterative PCR enrichment processes and the limited structural diversity of the initial library, which seriously restrict their real-world applications. To overcome these limitations, the in silico base mutagenesis post-SELEX strategy based on the low Gibbs free energy (ΔG) and genetic algorithm was developed for the optimization of the interferon-gamma aptamer (B1-4). In the process of evolution, new sequences were created and the aptamer candidates with low ΔG values and advanced structures were produced. After five rounds of selection, systematic studies revealed that the affinity of the newly developed evolutionary aptamer (M5-5) was roughly 10-fold higher than that of the parent aptamer (B1-4), and an aptasensor detection system with a limit-of-detection (LOD) value of 3.17 nM was established based on the evolutionary aptamer. The proposed approach provided an efficient strategy to improve the aptamer with low energy and a high binding ability, and the good analytical utility thereof. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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9 pages, 1099 KiB

Open AccessArticle

A Simple Structure-Switch Aptasensor Using Label-Free Aptamer for Fluorescence Detection of Aflatoxin B1

by Chao Wang, Hao Yu and Qiang Zhao

Molecules 2022, 27(13), 4257; https://doi.org/10.3390/molecules27134257 - 01 Jul 2022

Cited by 7 | Viewed by 1923

Abstract

Aflatoxin B1 (AFB1) is one of the mycotoxins produced by Aspergillus flavus and Aspergillus parasiticus, and it causes contamination in foods and great risk to human health. Simple sensitive detection of AFB1 is important and demanded for food safety and quality control. [...] Read more.

Aflatoxin B1 (AFB1) is one of the mycotoxins produced by Aspergillus flavus and Aspergillus parasiticus, and it causes contamination in foods and great risk to human health. Simple sensitive detection of AFB1 is important and demanded for food safety and quality control. Aptamers can specifically bind to targets with high affinity, showing advantages in affinity assays and biosensors. We reported an aptamer structure-switch for fluorescent detection of aflatoxin B1 (AFB1), using a label-free aptamer, a fluorescein (FAM)-labeled complementary strand (FDNA), and a quencher (BHQ1)-labeled complementary strand (QDNA). When AFB1 is absent, these three strands assemble into a duplex DNA structure through DNA hybridization, making FAM close to BHQ1, and fluorescence quenching occurs. In the presence of AFB1, the aptamer binds with AFB1, instead of hybridizing with QDNA. Thus, FAM is apart from BHQ1, and fluorescence increases with the addition of AFB1. This assay allowed detection of AFB1 with a detection limit of 61 pM AFB1 and a dynamic concentration range of 61 pM to 4 μM. This aptamer-based method enabled detection of AFB1 in complex sample matrix (e.g., beer and corn flour samples). Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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10 pages, 1179 KiB

Open AccessArticle

The Ability of Nuclease-Resistant RNA Aptamer against Streptococcus suis Serotype 2, Strain P1/7 to Reduce Biofilm Formation In Vitro

by Apinyapat Matchawong, Chatchawan Srisawat, Sirikwan Sangboonruang and Chayada Sitthidet Tharinjaroen

Molecules 2022, 27(12), 3894; https://doi.org/10.3390/molecules27123894 - 17 Jun 2022

Cited by 5 | Viewed by 1824

Abstract

Streptococcus suis, a Gram-positive bacterium, is an important swine and human pathogen, with serotype 2 being the most prevalent strain found worldwide. Deafness, meningitis, and death (in severe cases) are observed in S. suis-infected cases. Development of the ligands that can [...] Read more.

Streptococcus suis, a Gram-positive bacterium, is an important swine and human pathogen, with serotype 2 being the most prevalent strain found worldwide. Deafness, meningitis, and death (in severe cases) are observed in S. suis-infected cases. Development of the ligands that can bind to S. suis with high affinity and specificity could be beneficial for the diagnosis and treatment of S. suis infection. Herein, the nuclease-resistant RNA aptamers based on 2′-fluoropyrimidine modification against S. suis serotype 2, strain P1/7, were established using the cell- Systematic Evolution of Ligands by Exponential enrichment (SELEX) technique. One of the aptamers, R8-su12, could bind to the S. suis target strain as well as other S. suis serotypes, i.e., 1, 1/2, 9, and 14, but not to other bacteria tested, i.e., S. pneumoniae ATCC 49619, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa ATCC 27853. Moreover, the R8-su12 RNA aptamer was also capable of inhibiting the biofilm formation of the S. suis target strain, making it potentially useful for the study of biofilm formation and the treatment of S. suis infection in humans and pigs in the future. Full article

(This article belongs to the Special Issue Aptamer Generation and Bioapplication)

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