Damira Kanayeva

Monkeypox (mpox) is an infectious disease caused by the mpox virus and can potentially lead to fatal outcomes. It resembles infections caused by viruses from other families, challenging identification. The pathogenesis, transmission, and clinical manifestations of mpox and other Orthopoxvirus species are similar due to their closely related genetic material. This review provides a comprehensive discussion of the roles of various proteins, including extracellular enveloped virus (EEV), intracellular mature virus (IMV), and profilin-like proteins of mpox. It also highlights recent diagnostic techniques based on these proteins to detect this infection rapidly.

The global pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused a threat to public health and a worldwide crisis. This raised the need for quick, effective, and sensitive detection tools to prevent the rapid transmission rate of the infection. Therefore, this study aimed to develop an electrochemical impedance spectroscopy (EIS)-based aptasensor employing an interdigitated gold electrode (IDE) to detect SARS-CoV-2 Spike (S) glycoprotein and viral particles. This allowed us to sensitively detect SARS-CoV-2 S glycoprotein with a limit of detection (LOD) of 0.4 pg/mL in a buffer solution and to obtain a linear increase for concentrations between 0.2 to 0.8 pg/mL with high specificity. The proposed aptasensor also showed a good sensitivity towards the heat-inactivated SARS-CoV-2 variants in a buffer solution, where the Delta, Wuhan, and Alpha variants were captured at a viral titer of 6.45 ± 0.16 × 10³ TCID₅₀/mL, 6.20 × 10⁴ TCID₅₀/mL, and 5.32 ± 0.13 × 10² TCID₅₀/mL, respectively. Furthermore, the detection of SARS-CoV-2 performed in a spiked human nasal fluid provided an LOD of 6.45 ± 0.16 × 10³ TCID₅₀/mL for the Delta variant in a 50 µL sample and a detection time of less than 25 min. Atomic force microscopy images complemented the EIS results in this study, revealing that the surface roughness of the IDE after each modification step increased, which indicates that the target was successfully captured. This label-free EIS-based aptasensor has promising potential for the rapid detection of SARS-CoV-2 in complex clinical samples.

One of the major causes of a drastically shorter life expectancy and one of the most prevalent diseases in the world today is cancer. Given the data on the rise in cancer cases throughout the world, it is obvious that, despite the diagnostic techniques currently being used, there is a pressing need to develop precise and sensitive techniques for early diagnosis of the disease. A high degree of affinity and specificity towards particular targets is maintained by the short nucleic acid molecules known as aptamers. Aptamers outperform antibodies due to their unique benefits, such as their simplicity in synthesis and modification, lack of toxicity, and long-term stability. Utilizing an accurate recognition element and a robust signal transduction mechanism, molecular diagnostics can be extremely sensitive and specific. In this study, development of new single-stranded DNA aptamers against CEA for use in cancer diagnostics was accomplished using SELEX and NGS methods. As a result of 12 iterative SELEX rounds, nine aptamer candidates against CEA were developed. NGS comparative analysis revealed that round twelve had an enriched number of aptamers that were specifically bound, as opposed to round eight. Among the selected nine sequences characterized by bioinformatics analysis and ELONA, an aptamer sequence with the highest specificity and affinity for the target protein was identified and further examined. Aptamer sequence (6) was screened in a concentration-dependent assay, specificity analysis was performed, and its potential secondary and tertiary structures were predicted, which enabled us to test one of the possible putative interactions with CEA. Finally, aptamer sequence (6) labelled with a Cy5 fluorescent tag was used in confocal microscopy to observe its binding towards the CEA expressed in HT-29 human colon adenocarcinoma cell line.

The exponential spread of COVID-19 has prompted the need to develop a simple and sensitive diagnostic tool. Aptamer-based detection assays like ELONA are promising since they are inexpensive and sensitive. Aptamers have advantages over antibodies in wide modification, small size, in vitro selection, and stability under stringent conditions, which aid in scalable and reliable detection. In this work, we used aptamers against SARS-CoV-2 RBD S protein to design a simple and sensitive ELONA detection tool. Screening CoV2-RBD-1C and CoV2-RBD-4C aptamers and optimizing assay conditions led to the development of a direct ELONA that can detect SARS-CoV-2 RBD S glycoprotein in buffer solution and 0.1 % human nasal fluid with a detection limit of 2.16 ng/mL and 1.02 ng/mL, respectively. We detected inactivated Alpha, Wuhan, and Delta variants of SARS-CoV-2 with the detection limit of 3.73, 5.72, and 6.02 TCID₅₀/mL, respectively. Using the two aptamers as capture and reporter elements, we designed a more sensitive sandwich assay to identify the three SARS-CoV-2 variants employed in this research. As predicted, a lower detection limit was obtained. Sandwich assay LOD was 2.31 TCID₅₀/mL for Alpha, 1.15 TCID₅₀/mL for Wuhan, and 2.96 TCID₅₀/mL for Delta. The sensitivity of sandwich ELONA was validated using Alpha and Wuhan variants spiked in 0.1% human nasal fluid sample condition and were detected in 1.41 and 1.79 TCID₅₀/mL LOD, respectively. SEM was used to visualize the presence of viral particles in the Delta variant sample. The effective detection of SARS-CoV-2 in this study confirms the potential of our aptamer-based technique as a screening tool.

To date, Ag-based nanomaterials have demonstrated a high potential to overcome antibiotic resistance issues. However, bare Ag nanomaterials are prone to agglomeration in the biological environment, which results in a loss of antibacterial activity over time. Furthermore, it is still challenging to collect small-sized Ag nanomaterials right after the synthesis process. In this study, spherical-shaped Ag nanoparticles (NPs) (~6–10 nm) were attached on the surface of cetyltrimethylammonium bromide (CTAB)-loaded mesoporous silica nanoparticles (MSNs) (~100–110 nm). Antibacterial activity tests suggested that the obtained nanocomposite can be used as a highly efficient antibacterial agent against both Gram-negative and Gram-positive bacterial strains. The minimum inhibitory concentration (MIC) recalculated to pure Ag weight in nanocomposite was found to be ~1.84 µg/mL (for Escherichia coli) and ~0.92 µg/mL (for Staphylococcus aureus)—significantly smaller compared to values reported to date. The improved antibacterial activity of the prepared nanocomposite can be attributed to the even distribution of non-aggregated Ag NPs per volume unit and the presence of CTAB in the nanocomposite pores.

Chemically modified metal surfaces have been used to recognize and capture specific cell types and biomolecules. In this work, stainless steel wires were functionalized with aptamers against breast cancer stem cell markers. Stainless steel wires were first electropolished and silanized via electrodeposition. Aptamers were then attached to the silanized surface through a cross-linker. The functionalized wires were able to capture the target cells in an in vitro test. During surface modification steps, wires were analyzed by atomic force microscopy, cyclic voltammetry, scanning electron and fluorescence microscopy to determine their surface composition and morphology. Optimized conditions of silanization (applied potential, solution pH, heat treatment temperature) for obtaining an aptamer-functionalized wire were determined in this work together with the use of several surface characterization techniques suitable for small-sized and circular wires. These modified wires have potential applications for the in vivo capture of target cells in blood flow, since their small size allows their insertion as standard guidewires in biomedical devices.

This work presents experimental results on detection of Mycobacterium tuberculosis secreted protein MPT64 using an interdigitated electrode (IDE) which acts as a platform for capturing an immunogenic protein and an electrochemical impedance spectroscopy (EIS) as a detection technique. The assay involves a special receptor, single stranded DNA (ssDNA) aptamer, which specifically recognizes MPT64 protein. The ssDNA immobilization on IDE was based on a co-adsorbent immobilization at an optimized ratio of a 1/100 HS-(CH₆)₆-OP(O)₂O-(CH₂CH₂O)₆-5′-TTTTT-aptamer-3′/6-mercaptohexanol. The optimal sample incubation time required for a signal generation on an aptamer modified IDE was found to be at a range of 15–20 min. Atomic Force Microscopy (AFM) results confirmed a possible formation of an aptamer - MPT64 complex with a 20 nm roughness on the IDE surface vs. 4.5 nm roughness for the IDE modified with the aptamer only. A limit of detection for the EIS aptasensor based on an IDE for the detection of MPT64 in measurement buffer was 4.1 fM. The developed EIS aptasensor was evaluated on both serum and sputum clinical samples from the same TB (−) and TB (+) patients having a specificity and sensitivity for the sputum sample analysis 100% and 76.47%, respectively, and for the serum sample analysis 100% and 88.24%, respectively. The developed aptasensor presents a sensitive method for the TB diagnosis with the fast detection time.