Screening of broad-spectrum aptamer and development of electrochemical aptasensor for simultaneous detection of penicillin antibiotics in milk.

Penicillin antibiotics (PENs) play an important role in killing pathogenic bacteria. However, the residues of various penicillin antibiotics in milk gradually accumulate in the human body with the increase of milk intake, which causes direct harm to the human body. Aptamers can be used as recognition element of sensors. It is great significance to use broad-spectrum aptamers for simultaneous detection of PENs. In this study, we reported the screening and identification of DNA aptamers for PENs. The aptamers were screened by graphene oxide-systematic evolution of ligands by exponential enrichment (GO-SELEX). The broad-spectrum aptamers with high affinity and specificity were successfully obtained after 13 rounds of screening. The affinity and specificity of candidate aptamers were analyzed by a GO fluorescence competition method. Further sequence analysis revealed that a truncated 47 nt aptamer (P-11-1) had a higher affinity than the original 79 nt aptamer. The truncated aptamer P-11-1 was used as a recognition element, and an electrochemical aptasensor was prepared using gold nanoparticles (AuNPs) combined with ferroferric oxide-multi walled carbon nanotube (Fe3O4-MWCNTs) complex. The results showed that the developed aptasensor achieved the simultaneous detection of PENs in milk samples across a concentration range of 2 nM-10,000 nM, achieving a limit of detection of 0.667 nM. This methodology provided a simple and sensitive new thinking for antibiotic multi-residue detection.

Paper-based biosensors relying on core biological immune scaffolds for the detection of procymidone in vegetables.

In order to achieve a highly sensitive detection of procymidone in vegetables, three paper-based biosensors based on a core biological immune scaffold (CBIS) were developed, which were time-resolved fluorescence immunochromatography strips with Europium (III) oxide (Eu-TRFICS). Goat anti-mouse IgG and europium oxide time-resolved fluorescent microspheres formed secondary fluorescent probes. CBIS was formed by secondary fluorescent probes and procymidone monoclonal antibody (PCM-Ab). The first type of Eu-TRFICS (Eu-TRFICS-(1)) fixed secondary fluorescent probes on a conjugate pad, and PCM-Ab was mixed with a sample solution. The second type of Eu-TRFICS (Eu-TRFICS-(2)) fixed CBIS on the conjugate pad. The third type of Eu-TRFICS (Eu-TRFICS-(3)) was directly mixed CBIS with the sample solution. They solved the problems of steric hindrance of antibody labeling, insufficient exposure of antigen recognition region and easy loss of activity in traditional methods. They realized multi-dimensional labeling and directional coupling. They replaced the loss of antibody activity. And the three types of Eu-TRFICS were compared, among which Eu-TRFICS-(1) was the best detection choice. Antibody usage was reduced by 25% and sensitivity was increased by 3 times. Its detection range was 1-800 ng/mL, the limit of detection (LOD) was 0.12 ng/mL with the visible LOD (vLOD) of 5 ng/mL.

Novel Electrochemical Aptasensor Based on Ordered Mesoporous Carbon/2D Ti3C2 MXene as Nanocarrier for Simultaneous Detection of Aminoglycoside Antibiotics in Milk.

Herein, a novel electrochemical aptasensor using a broad-spectrum aptamer as a biorecognition element was constructed based on a screen-printed carbon electrode (SPCE) for simultaneous detection of aminoglycoside antibiotics (AAs). The ordered mesoporous carbon (OMC) was firstly modified on 2D Ti3C2 MXene. The addition of OMC not only effectively improved the stability of the aptasensor, but also prevented the stacking of Ti3C2 sheets, which formed a good current passage for signal amplification. The prepared OMC@Ti3C2 MXene functioned as a nanocarrier to accommodate considerable aptamers. In the presence of AAs, the transport of electron charge on SPCE surface was influenced by the bio-chemical reactions of the aptamer and AAs, generating a significant decline in the differential pulse voltammetry (DPV) signals. The proposed aptasensor presented a wide linear range and the detection limit was 3.51 nM. Moreover, the aptasensor, with satisfactory stability, reproducibility and specificity, was successfully employed to detect the multi-residuals of AAs in milk. This work provided a novel strategy for monitoring AAs in milk.

Aptamers against Pathogenic Bacteria: Selection Strategies and Apta-assay/Aptasensor Application for Food Safety.

Pathogenic bacteria are primarily kinds of detrimental agents that cause mankind illness via contaminated food with traits of multiple types, universality, and low content. In view of the detection demands for rapidity, aptamer recognition factors emerged as a substitution for antibodies, which are short single strands of nucleic acid selected via in vitro. They display certain superiorities over antibodies, such as preferable stability, liable modification, and cost-efficiency. Taking advantage of the situation, numerous aptamers against pathogenic bacteria have been successfully selected and applied, yet there are still restrictions on commercial availability. In this review, the strategies/approaches to key sections in pathogen aptamers SELEX and post-SELEX are summarized and sorted out. Recently, optical, electrochemical, and piezoelectric aptamer-based assays or sensors dedicated to pathogen detection have been critically reviewed. Ultimately, the existing challenges and future trends in this field are proposed to further promote development prospects.

Non-immobilized GO-SELEX of aptamers for label-free detection of thiamethoxam in vegetables.

Due to the massive use of thiamethoxam (TMX) pesticide and the accumulated potential hazards exposure, the detection of TMX is of great significance to food and ecological safety. In this study, aptamers with affinity for TMX were obtained through graphene oxide assisted systematic evolution of ligands by exponential enrichment (GO-SELEX). After 9 rounds of positive and counter selection, 5 candidate sequences were obtained, among which seq.20 had the highest affinity for TMX, and its dissociation constant (Kd) was 210.47 ± 79.37 nM. Then, the aptamer was further truncated based on structural analysis. The truncated aptamers (seq.20-1, seq.20-2) exhibited higher affinity (Kd = 118.34 ± 13.85 nM, Kd = 123.35 ± 29.80 nM), which seq.20-2 had only 37 bases. Furthermore, circular dichroism spectroscopy showed that TMX induced the conformation of aptamer from B-form structure to hairpin structure, and then formed a stable TMX-ssDNA complex. Finally, the truncated aptamer (seq.20-2) and the original aptamer (seq.20) were used as recognition elements to construct colorimetric aptasensors based on gold nanoparticles for the detection of TMX. It was found that the sensitivity of the former (LOD = 1.67 ± 0.12 nM, S/N = 3) was better than that of the latter (LOD = 3.33 ± 0.23 nM, S/N = 3). Feasibility of truncated aptamer as recognition element in the detection of TMX in vegetable samples was preliminarily verified.

Recent advances in aptamer-based sensors for aminoglycoside antibiotics detection and their applications.

Aminoglycoside antibiotics (AAs) have been extensively applied in medical field and animal husbandry owing to desirable broad-spectrum antibacterial activity. Excessive AAs residues in the environment can be accumulated in human body through food chain and cause detrimental effect on human health. The establishment of highly specific, simple and sensitive detection methods for monitoring AAs residues is highly in demand. Aptasensor using aptamer as the biological recognition element is the efficient and promising sensing method for detection of AAs. In this review, we have made a summary of specific aptamers sequences against AAs. Subsequently, we provide a systematical and comprehensive overview of modern techniques in aptasensors for detection of AAs according to optical aptasensors as well as electrochemical aptasensors and further summarize their advantages and disadvantages to compare their applications. In addition, we present an overview of practical applications of aptasensors in sample detection of AAs. Moreover, the current challenges and future trends in this field are also included to reveal a promising perspective for developing novel aptasensors for AAs.