Biparatopic anti-PCSK9 antibody enhances the LDL-uptake in HepG2 cells.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) has emerged as a promising therapeutic target to reduce lipids. In 2020, we reported a chimeric camelid-human heavy chain antibody VHH-B11-Fc targeting PCSK9. Recently, it was verified that VHH-B11 binds one linear epitope in the PCSK9 hinge region. To enhance its druggability, we have developed a novel biparatopic B11-H2-Fc Ab herein. Thereinto, surface plasmon resonance (SPR) confirmed the epitope differences in binding-PCSK9 among VHH-B11, VHH-H2 and the approved Repatha. Additionally, SPR revealed the B11-H2-Fc exhibits an avidity of approximately 0.036 nM for PCSK9, representing a considerable increase compared to VHH-B11-Fc (~ 0.69 nM). Moreover, we found the Repatha and B11-H2-Fc exhibited > 95% PCSK9 inhibition efficiency compared to approximately 48% for the VHH-Fc at 7.4 nM (P < 0.0005). Further, we verified its biological activity using the human hepatoma cells G2 model, where the B11-H2-Fc exhibited almost 100% efficiency in PCSK9 inhibition at only 0.75 µM. The immunoblotting results of low-density lipoprotein cholesterol (LDL-c) uptake assay also demonstrated the excellent performance of B11-H2-Fc on recovering the LDL-c receptor (LDLR), as strong as the Repatha (P > 0.05). These findings provide the first evidence of the efficacy of a novel Ab targeting PCSK9 in the field of lipid-lowering drugs.

Early determination of potential critical quality attributes of therapeutic antibodies in developability studies through surface plasmon resonance-based relative binding activity assessment.

Precise measurement of the binding activity changes of therapeutic antibodies is important to determine the potential critical quality attributes (CQAs) in developability assessment at the early stage of antibody development. Here, we report a surface plasmon resonance (SPR)-based relative binding activity method, which incorporates both binding affinity and binding response and allows us to determine relative binding activity of antibodies with high accuracy and precision. We applied the SPR-based relative binding activity method in multiple forced degradation studies of antibody developability assessment. The current developability assessment strategy provided comprehensive, precise characterization of antibody binding activity in the stability studies, enabling us to perform correlation analysis and establish the structure-function relationship between relative binding activity and quality attributes. The impact of a given quality attribute on binding activity could be confidently determined without isolating antibody variants. We identified several potential CQAs, including Asp isomerization, Asn deamidation, and fragmentation. Some potential CQAs affected binding affinity of antibody and resulted in a reduction of binding activity. Certain potential CQAs impaired antibody binding to antigen and led to a loss of binding activity. A few potential CQAs could influence both binding affinity and binding response and cause a substantial decrease in antibody binding activity. Specifically, we identified low abundance Asn33 deamidation in the light chain complementarity-determining region as a potential CQA, in which all the stressed antibody samples showed Asn33 deamidation abundances ranging from 4.2% to 27.5% and a mild binding affinity change from 1.76 nM to 2.16 nM.

Target-triggered assembly of plasmon resonance nanostructures for quantitative detection of lncRNA in liver cancer cells via surface enhanced Raman spectroscopy.

Long-stranded non-coding RNAs (lncRNA) have important roles in disease as transcriptional regulators, mRNA processing regulators and protein synthesis factors. However, traditional methods for detecting lncRNA are time-consuming and labor-intensive, and the functions of lncRNA are still being explored. Here, we present a surface enhanced Raman spectroscopy (SERS) based biosensor for the detection of lncRNA associated with liver cancer (LC) as well as in situ cellular imaging. Using the dual SERS probes, quantitative detection of lncRNA (DAPK1-215) can be achieved with an ultra-low detection limit of 952 aM by the target-triggered assembly of core-satellite nanostructures. And the reliability of this assay can be further improved with the R2 value of 0.9923 by an internal standard probe that enables the signal dynamic calibration. Meanwhile, the high expression of DAPK1-215 mainly distributed in the cytoplasm was observed in LC cells compared with the normal ones using the SERS imaging method. Moreover, results of cellular function assays showed that DAPK1-215 promoted the migration and invasion of LC by significantly reducing the expression of the structural domain of death associated protein kinase. The development of this biosensor based on SERS can provide a sensitive and specific method for exploring the expression of lncRNA that would be a potential biomarker for the screening of LC.

Corrosive Pseudomonas aeruginosa detection by measuring pyocyanin with a lab-on-fiber optical surface plasmon resonance biosensor in aquatic environments.

Oceanic facilities and equipment corrosion present considerable economic and safety concerns, predominantly due to microbial corrosion. Early detection of corrosive microbes is pivotal for effective monitoring and prevention. Yet, traditional detection methods often lack specificity, require extensive processing time, and yield inaccurate results. Hence, the need for an efficient real-time corrosive microbe monitoring technology is evident. Pseudomonas aeruginosa, a widely distributed microorganism in aquatic environments, utilizes its production of quinone-like compounds, specifically pyocyanin (PYO), to corrode metals. Here, we report a novel fiber optic surface plasmon resonance (SPR) sensor modified by the C-terminal of BrlR protein (BrlR-C), which is a specific receptor of PYO molecule, to detect P. aeruginosa in aquatic environments. The results showed that the sensor had a good ability to recognize PYO in the concentration range of 0-1 µg/mL, and showed excellent sensing performance in real-time monitoring the growth status of P. aeruginosa. With a strong selectivity of PYO, the sensor could clearly detect P. aeruginosa against other bacteria in seawater environment, and exhibited excellent anti-interference ability against variations in pH, temperature and pressure and other interfering substances. This study provides a useful tool for monitoring corrosive P. aeruginosa biofilm in aquatic environments, which is a first of its kind example that serves as a laboratory model for the application of fiber optic technology in real-world scenarios to monitoring biofilms in microbial corrosion and biofouling.

Objective-Free Ultrasensitive Biosensing on Large-Area Metamaterial Surfaces in the Near-IR.

Plasmonic metamaterials have opened new avenues in medical diagnostics. However, the transfer of the technology to the markets has been delayed due to multiple challenges. The need of bulky optics for signal reading from nanostructures patterned on submillimeter area limits the miniaturization of the devices. The use of objective-free optics can solve this problem, which necessitates large area patterning of the nanostructures. In this work, we utilize laser interference lithography (LIL) to pattern nanodisc-shaped metamaterial absorber nanoantennas over a large area (4 cm2) within minutes. The introduction of a sacrificial layer during the fabrication process enables an inverted hole profile and a well-controlled liftoff, which ensures perfectly defined uniform nanopatterning almost with no defects. Furthermore, we use a macroscopic reflection probe for optical characterization in the near-IR, including the detection of the binding kinematics of immunologically relevant proteins. We show that the photonic quality of the plasmonic nanoantennas commensurates with electron-beam-lithography-fabricated ones over the whole area. The refractive index sensitivity of the LIL-fabricated metasurface is determined as 685 nm per refractive index unit, which demonstrates ultrasensitive detection. Moreover, the fabricated surfaces can be used multiple times for biosensing without losing their optical quality. The combination of rapid and large area nanofabrication with a simple optical reading not only simplifies the detection process but also makes the biosensors more environmentally friendly and cost-effective. Therefore, the improvements provided in this work will empower researchers and industries for accurate and real-time analysis of biological systems.

Uricase biofunctionalized plasmonic sensor for uric acid detection with APTES-modified gold nanotopping.

Current uric acid detection methodologies lack the requisite sensitivity and selectivity for point-of-care applications. Plasmonic sensors, while promising, demand refinement for improved performance. This work introduces a biofunctionalized sensor predicated on surface plasmon resonance to quantify uric acid within physiologically relevant concentration ranges. The sensor employs the covalent immobilization of uricase enzyme using 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-Hydroxysuccinimide (NHS) crosslinking agents, ensuring the durable adherence of the enzyme onto the sensor probe. Characterization through atomic force microscopy and Fourier transform infrared spectroscopy validate surface alterations. The Langmuir adsorption isotherm model elucidates binding kinetics, revealing a sensor binding affinity of 298.83 (mg/dL)-1, and a maximum adsorption capacity of approximately 1.0751°. The biofunctionalized sensor exhibits a sensitivity of 0.0755°/(mg/dL), a linear correlation coefficient of 0.8313, and a limit of detection of 0.095 mg/dL. Selectivity tests against potentially competing interferents like glucose, ascorbic acid, urea, D-cystine, and creatinine showcase a significant resonance angle shift of 1.1135° for uric acid compared to 0.1853° for interferents at the same concentration. Significantly, at a low uric acid concentration of 0.5 mg/dL, a distinct shift of 0.3706° was observed, setting it apart from the lower values noticed at higher concentrations for all typical interferent samples. The uricase enzyme significantly enhances plasmonic sensors for uric acid detection, showcasing a seamless integration of optical principles and biological recognition elements. These sensors hold promise as vital tools in clinical and point-of-care settings, offering transformative potential in biosensing technologies and the potential to revolutionize healthcare outcomes in biomedicine.

A new di-recognition and di-functional nanosurface aptamer molecularly imprinted polymer probe for trace glyphosate with SERS/RRS/Abs trimode technique.

A new di-recognition nitrogen-doped carbon dot nanosurface aptamer molecularly imprinted polymer (CDNAg@MIPApt) nanocatalytic di-functional probe was prepared by microwave irradiation. The probe was utilized nitrogen-doped silver carbon dots (CDNAg) as the matrix, glyphosate (Gly) as the template molecule, α-methyl acrylate as the monomer, ethylene glycol dimethacrylate as the cross-linker, and aptamer as the biorecognition element. It could not only recognize Gly but also exhibits catalytic amplification function. It was found that CDNAg@MIPApt catalyzed the redox reaction of polyethylene glycol 400 (PEG400)-AgNO3 to generate silver nanoparticles (AgNPs). The AgNPs indicator component exhibit the effects of surface-enhanced Raman scattering (SERS), resonance Rayleigh scattering (RRS) and surface plasmon resonance absorption (Abs). In the presence of Gly, it binds to the surface imprinted site of CDNAg@MIPApt, to reduce AgNPs generation due to the catalytic activity of CDNAg@MIPApt decreasing. Thus, the SERS/RRS/Abs signal values decreased linearly. The linear ranges of SERS/RRS/Abs assay were 0.1-2.5 nM, 0.25-2.75 nM and 0.5-5 nM respectively. The detection limits were 0.034 nM, 0.071 nM and 0.18 nM Gly.

A cavity induced mode hybridization plasmonic sensor for portable detection of exosomes.

Exosomes have been considered as promising biomarkers for cancer diagnosis due to their abundant information from originating cells. However, sensitive and reliable detection of exosomes is still facing technically challenges due to the lack of a sensing platform with high sensitivity and reproducibility. To address the challenges, here we propose a portable surface plasmon resonance (SPR) sensing of exosomes with a three-layer Au mirror/SiO2 spacer/Au nanohole sensor, fabricated by an economical polystyrene nanosphere self-assembly method. The SiO2 spacer can act as an optical cavity and induce mode hybridization, leading to excellent optimization of both sensitivity and full width at half maximum compared with normal single layer Au nanohole sensors. When modified with CD63 or EpCAM aptamers, a detection of limit (LOD) of as low as 600 particles/µL was achieved. The sensors showed good capability to distinguish between non-tumor derived L02 exosomes and tumor derived HepG2 exosomes. Additionally, high reproducibility was also achieved in detection of artificial serum samples with RSD as low as 2%, making it feasible for clinical applications. This mode hybridization plasmonic sensor provides an effective approach to optimize the detection sensitivity of exosomes, pushing SPR sensing one step further towards cancer diagnosis.

Advances in Nanoplasmonic Biosensors: Optimizing Performance for Exosome Detection Applications.

The development of sensitive and specific exosome detection tools is essential because they are believed to provide specific information that is important for early detection, screening, diagnosis, and monitoring of cancer. Among the many detection tools, surface-plasmon resonance (SPR) biosensors are analytical devices that offer advantages in sensitivity and detection speed, thereby making the sample-analysis process faster and more accurate. In addition, the penetration depth of the SPR biosensor, which is <300 nm, is comparable to the size of the exosome, making the SPR biosensor ideal for use in exosome research. On the other hand, another type of nanoplasmonic sensor, namely a localized surface-plasmon resonance (LSPR) biosensor, has a shorter penetration depth of around 6 nm. Structural optimization through the addition of supporting layers and gap control between particles is needed to strengthen the surface-plasmon field. This paper summarizes the progress of the development of SPR and LSPR biosensors for detecting exosomes. Techniques in signal amplification from two sensors will be discussed. There are three main parts to this paper. The first two parts will focus on reviewing the working principles of each sensor and introducing several methods that can be used to isolate exosomes. This article will close by explaining the various sensor systems that have been developed and the optimizations carried out to obtain sensors with better performance. To illustrate the performance improvements in each sensor system discussed, the parameters highlighted include the detection limit, dynamic range, and sensitivity.

Expression and purification of human neutrophil proteinase 3 from insect cells and characterization of ligand binding.

Neutrophil proteinase 3 (PR3) is an important drug target for inflammatory lung diseases such as chronic obstructive pulmonary disease and cystic fibrosis. Drug discovery efforts targeting PR3 require active enzyme for in vitro characterization, such as inhibitor screening, enzymatic assays, and structural studies. Recombinant expression of active PR3 overcomes the need for enzyme supplies from human blood and in addition allows studies on the influence of mutations on enzyme activity and ligand binding. Here, we report the expression of recombinant PR3 (rPR3) using a baculovirus expression system. The purification and activation process described resulted in highly pure and active PR3. The activity of rPR3 in the presence of commercially available inhibitors was compared with human PR3 by using a fluorescence-based enzymatic assay. Purified rPR3 had comparable activity to the native human enzyme, thus being a suitable alternative for enzymatic studies in vitro. Further, we established a surface plasmon resonance-based assay to determine binding affinities and kinetics of PR3 ligands. These methods provide valuable tools for early drug discovery aiming towards treatment of lung inflammation.

Electrochemical and Plasmonic Detection of Myocardial Infarction Using Microfluidic Biochip Incorporated with Mesoporous Nanoscaffolds.

This paper reports a microfluidic device for the electrochemical and plasmonic detection of cardiac myoglobin (cMb) and cardiac troponin I (cTnI) with noticeable limits of detection (LoD) as low as a few picograms per milliliter (pg/mL) ranges, achieved in a short detection time. The device features two working electrodes, each with a mesoporous Ni3V2O8 nanoscaffold grafted with reduced graphene oxide (rGO) that improves the interaction of diffusing analyte molecules with the sensing surface by providing a high surface area and reaction kinetics. Electrochemical studies reveal sensitivities as high as 9.68 µA ng/mL and a LoD of 2.0 pg/mL for cTnI, and 8.98 µA ng/mL and 4.7 pg/mL for cMb. Additionally, the surface plasmon resonance (SPR) studies demonstrate a low-level LoD of 8.8 pg/mL for cMb and 7.3 pg/mL for cTnI. The dual-modality sensor enables dynamic tracking of kinetic antigen-antibody interactions during sensing, self-verification through providing signals of two modes, and reduced false readout. This study demonstrates the complementary nature of the electrochemical and SPR modes in biosensing, with the electrochemical mode being highly sensitive and the SPR mode providing superior tracking of molecular recognition behaviors. The presented sensor represents a significant innovation in cardiovascular disease management and can be applied to monitor other clinically important biomolecules.

Plasmon resonances of GZO core-Ag shell nanospheres, nanorods, and nanodisks for biosensing and biomedical applications in near-infrared biological windows I and II.

There is currently a great deal of interest in realizing localized surface plasmon resonances (LSPRs) in two distinct windows in the near-infrared (NIR) spectrum for in vivo biosensing and medical applications, the biological window (BW) I and II (BW I, 700-900 nm; BW II, 1000-1700 nm). This study aims to demonstrate that LSPRs of Ga-doped ZnO (GZO) core-silver (Ag) shell structures exhibit promising features for biological applications in the NIR BW I and II. Here, we study three different shapes for nanoshells: the core-shell nanosphere, nanorod, and nanodisk. In the calculation of the optical response of these nanoshells, an effective medium approach is first used to reduce the dielectric function of a nanoshell to that of an equivalent homogenous NP with an effective dielectric function. Then, the LSPR spectra of nanoshells are calculated using the modified long-wavelength approximation (MLWA), which corrects the polarizability of the equivalent NP as obtained by Gans theory. Through numerical investigations, we examine the impacts of the core and shell sizes of the proposed nanoshells as well as the medium refractive index on the position and line width of the plasmon resonance peaks. It is shown that the plasmon resonances of the three proposed nanoshells exhibit astonishing resonance tunability in the NIR region by varying their geometrical parameters. Specifically, the improved spectrum characteristics and tunability of its plasmon resonances make the GZO-Ag nanosphere a more viable platform for NIR applications than the spherical metal colloid. Furthermore, we demonstrate that the sensitivity and figure of merit (FOM) of the plasmon resonances may be significantly increased by using GZO-Ag nanorods and nanodisks in place of GZO-Ag nanospheres. It is found that the optical properties of the transverse plasmon resonance of the GZO-Ag nanodisk are superior to all plasmon resonances produced by the GZO-Ag nanorods and GZO-Ag nanospheres in terms of sensitivity and FOM. The FOM of the transverse plasmon mode of the GZO-Ag nanodisk is almost two orders of magnitude higher than that of the longitudinal and transverse plasmon modes of the GZO-Ag nanorod in BW I and BW II. And it is 1.5 and 2 times higher than the plasmon resonance FOM of GZO-Ag nanospheres in BW I and BW II, respectively.

Trace-Amount Detection of Chiral Molecules Based on Plasmonic Racemic Arrays Fabricated via Direct Laser Writing.

Superchiral fields, supported by chiral plasmonic structures, have shown outstanding performance for chiral molecule sensing via enhanced chiral light-matter interaction. However, this sensing capability cannot fully reveal the chiral origin of the molecules as the chiroptic response of the molecules is intertwined with the chiroptic response of the chiral plasmonic nanostructures, which can potentially be excluded by using a plasmonic racemic mixture. Such a plasmonic racemic mixture is not easily attainable, as it normally requires complex fabrication and expensive instrumentation, whose structural fineness is limited by the fabrication precision. Here, we demonstrate trace-amount chiral molecule detection with plasmonic racemic arrays fabricated by direct laser writing with vector beams, which is facile, cost-effective, and highly controllable. The racemic arrays present no inherent circular differential scattering but a large local superchiral field, which reflects the intrinsic chiral features of the chiral molecules. They are further applied to discriminate enantiomers of phenylalanine with a limit of detection (LOD) of 10.0 ± 2.8 µM, which is an order of magnitude smaller than the LOD of conventional circular dichroism spectroscopy. The strong local superchiral field provided by the plasmonic racemic arrays enlightens the design of a superior sensing platform, which holds promising applications for biomedical detection and enantioselective drug development.

Dual-Mode Comb Plasmonic Optical Fiber Sensing.

Surface plasmon (SP) excitation in metal-coated tilted fiber Bragg gratings (TFBGs) has been a focal point for highly sensitive surface biosensing. Previous efforts focused on uniform metal layer deposition around the TFBG cross section and temperature self-compensation with the Bragg mode, requiring both careful control of the core-guided light polarization and interrogation over most of the C + L bands. To circumvent these two important practical limitations, we studied and developed an original platform based on partially coated TFBGs. The partial metal layer enables the generation of dual-comb resonances, encompassing highly sensitive (TM/EH mode families) and highly insensitive (TE/HE mode families) components in unpolarized transmission spectra. The interleaved comb of insensitive modes acts as wavelength and power references within the same spectral region as the SP-active modes. Despite reduced fabrication and measurement complexity, refractometric accuracy is not compromised through statistical averaging over seven individual resonances within a narrowband window of 10 nm. Consequently, measuring spectra over 60 nm is no longer needed to compensate for small temperature or power fluctuations. This sensing platform brings the following important practical assets: (1) a simpler fabrication process, (2) no need for polarization control, (3) limited bandwidth interrogation, and (4) maintained refractometric accuracy, which makes it a true game changer in the ever-growing plasmonic sensing domain.

A near-infrared plasmonic biosensor for detection of morphine and codeine in biological samples based on the end-to-end assembly of modified gold nanorods.

The analytical determination of opiates in biological samples is a critical mission and remains a challenge for almost all judicial and clinical drug testing panels due to their high abuse potential. Based on the high sensitivity of the longitudinal surface plasmon resonance (LSPR) peak of gold nanorods (AuNRs), we successfully developed a novel and simple refractive index sensing platform for detection of morphine (MOR) and codeine (COD) by means of 2-amino-5-mercapto-1,3,4-thiadiazole functionalized gold nanorods (AMTD-AuNRs) in aqueous solution, which is, to the best of our knowledge, the first report on the assay of MOR and COD using AuNRs. AMTD molecules strongly anchor onto the tips of AuNRs via the mercapto group and subsequent hydrogen-bonding interactions between AMTD and the analytes induced end-to-end chain assembly of AuNRs and a consequent decrease of the LSPR absorption band at 850 nm along with a bathochromic shift and emergence of a new hybridized plasmon mode at 1050 nm which was characterized using a Vis-NIR spectrophotometer. After systematic optimization, the absorbance ratio (A1050/A850) was proportional to the concentration of MOR in the ranges of 0.08-5 µM and 0.2-8 µM for COD without any significant effect from possible interferents. Furthermore, detection limits of 40 and 62 nM were achieved for MOR and COD, respectively, which are much lower than the cut-off level of 2000 ng mL-1 for opiates in urine samples set by the Substance and Abuse Mental Health Services Administration (SAMHSA). Eventually, as proof-of-applicability, human urine and blood serum samples spiked with MOR and COD were analyzed and excellent recoveries ranging from 94.4 to 108.9% were obtained, demonstrating the successful applicability of the designed refractive index probe in real biological specimens.

Use of Surface Plasmon Resonance Technique for Studies of Inter-domain Interactions in Ion Channels.

Ion channels are transmembrane proteins essential for cellular functions and are important drug targets. Surface plasmon resonance (SPR) is a powerful technique for investigating protein-protein and protein-small molecule ligand interactions. SPR has been underutilized for studies of ion channels, even though it could provide a wealth of information on the mechanisms of ion channel regulation and aid in ion channel drug discovery. Here we provide a detailed description of the use of SPR technology for investigating inter-domain interactions in KCNH potassium-selective and voltage-gated ion channels.

Applications of Surface Plasmon Resonance (SPR) to the Study of Diverse Protein-Ligand Interactions.

Functional characterization of enzymes/proteins requires determination of the binding affinity of small molecules or other biomolecules with the target proteins. Several available techniques, such as proteomics and drug discovery strategies, require a precise and high-throughput assay for rapid and reliable screening of potential candidates for further testing. Surface plasmon resonance (SPR), a well-established label-free technique, directly measures biomolecular affinities. SPR assays require immobilization of one interacting component (ligand) on a conductive metal (mostly gold or silver) and a continuous flow of solution containing potential binding partner (analyte) across the surface. The SPR phenomenon occurs when polarized light excites the electrons at the interface of the metal and the dielectric medium to generate electromagnetic waves that propagate parallel to the surface. Changes in the refractive index due to interaction between the ligand and analyte are measured by detecting the reflected light, providing real-time data on kinetics and specificity. A prominent use of SPR is identifying compounds in crude plant extracts that bind to specific molecules. Procedures that utilize SPR are becoming increasingly applicable outside the laboratory setting, and SPR imaging and localized SPR (LSPR) are cheaper and more portable alternative for in situ detection of plant or mammalian pathogens and drug discovery studies. LSPR, in particular, has the advantage of direct attachment to test tissues in live-plant studies. Here, we describe three protocols utilizing SPR-based assays for precise analysis of protein-ligand interactions. © 2024 Wiley Periodicals LLC. Basic Protocol 1: SPR comparison of binding affinities of viral reverse transcriptase polymorphisms Basic Protocol 2: SPR screening of crude plant extract for protein-binding agents Basic Protocol 3: Localized SPR-based antigen detection using antibody-conjugated gold nanoparticles.

Novel benzimidazole angiotensin receptor blockers with anti-SARS-CoV-2 activity equipotent to that of nirmatrelvir: computational and enzymatic studies.

BACKGROUND: Hypertension worsens outcomes in SARS-CoV-2 patients. Sartans, a type of antihypertensive angiotensin receptor blocker-(ARB), reduce COVID-19 morbidity and mortality by targeting angiotensin-converting enzyme-2 (ACE2). This study aimed to evaluate the antiviral and antihypertensive effects of nirmatrelvir, commercial sartans (candesartan, losartan, and losartan carboxylic (Exp3174)), and newly synthesized sartans (benzimidazole-N-biphenyl carboxyl (ACC519C) and benzimidazole-N-biphenyl tetrazole (ACC519T)), compared to nirmatrelvir, the antiviral component of Paxlovid. RESEARCH DESIGN AND METHODS: Surface plasmon resonance (SPR) and enzymatic studies assessed drug effects on ACE2. Antiviral abilities were tested with SARS-CoV-2-infected Vero E6 cells, and antihypertensive effects were evaluated using angiotensin II-contracted rabbit iliac arteries. RESULTS: Benzimidazole-based candesartan and ACC519C showed antiviral activity comparable to nirmatrelvir (95% inhibition). Imidazole-based losartan, Exp3174, and ACC519T were less potent (75%-80% and 50%, respectively), with Exp3174 being the least effective. SPR analysis indicated high sartans-ACE2 binding affinity. Candesartan and nirmatrelvir combined had greater inhibitory and cytopathic effects (3.96%) than individually (6.10% and 5.08%). ACE2 enzymatic assays showed varying effects of novel sartans on ACE2. ACC519T significantly reduced angiotensin II-mediated contraction, unlike nirmatrelvir and ACC519T(2). CONCLUSION: This study reports the discovery of a new class of benzimidazole-based sartans that significantly inhibit SARS-CoV-2, likely due to their interaction with ACE2.

Near-infrared-II responsive ovalbumin functionalized gold-genipin nanosystem cascading photo-immunotherapy of cancer.

Synergistic cancer therapies have attracted wide attention owing to their multi-mode tumor inhibition properties. Especially, photo-responsive photoimmunotherapy demonstrates an emerging cancer treatment paradigm that significantly improved treatment efficiency. Herein, near-infrared-II responsive ovalbumin functionalized Gold-Genipin nanosystem (Au-G-OVA NRs) was designed for immunotherapy and deep photothermal therapy of breast cancer. A facile synthesis method was employed to prepare the homogeneous Au nanorods (Au NRs) with good dispersion. The nanovaccine was developed further by the chemical cross-linking of Au-NRs, genipin and ovalbumin. The Au-G-OVA NRs outstanding aqueous solubility, and biocompatibility against normal and cancer cells. The designed NRs possessed enhanced localized surface plasmon resonance (LSPR) effect, which extended the NIR absorption in the second window, enabling promising photothermal properties. Moreover, genipin coating provided complimentary red fluorescent and prepared Au-G-OVA NRs showed significant intracellular encapsulation for efficient photoimmunotherapy outcomes. The designed nanosystem possessed deep photothermal therapy of breast cancer and 90% 4T1 cells were ablated by Au-G-OVA NRs (80µg ml-1concentration) after 1064 nm laser irradiation. In addition, Au-G-OVA NRs demonstrated outstanding vaccination phenomena by facilitating OVA delivery, antigen uptake, maturation of bone marrow dendritic cells, and cytokine IFN-γsecretion for tumor immunosurveillance. The aforementioned advantages permit the utilization of fluorescence imaging-guided photo-immunotherapy for cancers, demonstrating a straightforward approach for developing nanovaccines tailored to precise tumor treatment.

Screening and identification of DNA nucleic acid aptamers against F1 protein of Yersinia pestis using SELEX method.

BACKGROUND: Yersinia pestis is a bacterium that causes the disease plague. It has caused the deaths of many people throughout history. The bacterium possesses several virulence factors (pPla, pFra, and PYV). PFra plasmid encodes fraction 1 (F1) capsular antigen. F1 protein protects the bacterium against host immune cells through phagocytosis process. This protein is specific for Y. pestis. Many diagnostic techniques are based on molecular and serological detection and quantification of F1 protein in different food and clinical samples. Aptamers are small nucleic acid sequences that can act as specific ligands for many targets.This study, aimed to isolate the high-affinity ssDNA aptamers against F1 protein. METHODS AND RESULTS: In this study, SELEX was used as the main strategy in screening aptamers. Moreover, enzyme-linked aptamer sorbent assay (ELASA) and surface plasmon resonance (SPR) were used to determine the affinity and specificity of obtained aptamers to F1 protein. The analysis showed that among the obtained aptamers, the three aptamers of Yer 21, Yer 24, and Yer 25 were selected with a KD value of 1.344E - 7, 2.004E - 8, and 1.68E - 8 M, respectively. The limit of detection (LoD) was found to be 0.05, 0.076, and 0.033 µg/ml for Yer 21, Yer 24, and Yer 25, respectively. CONCLUSION: This study demonstrated that the synthesized aptamers could serve as effective tools for detecting and analyzing the F1 protein, indicating their potential value in future diagnostic applications.