Surface Binding Research Articles

Surface Functionalized Plasmonic Sensors for Uric Acid Detection With Gold-Graphene Stacked Nanocomposites

Abstract This study presented a surface-functionalized sensor probe using 3-aminopropyltriethoxysilane (APTES) self-assembled monolayers on a Kretschmann-configured plasmonic platform. The probe featured stacked nanocomposites of gold (via sputtering) and graphene quantum dots (GQD, via spin-coating) for highly sensitive and accurate uric acid (UA) detection within the physiological ranges. Characterization encompassed the field emission scanning electron microscopy for detailed imaging, energy-dispersive X-ray spectroscopy for elemental analysis, and Fourier transform infrared spectroscopy for molecular identification. Surface functionalization increased sensor sensitivity by 60.64%, achieving 0.0221 °/(mg/dL) for the gold-GQD probe and 0.035 5 °/(mg/dL) for the gold-APTES-GQD probe, with linear correlation coefficients of 0.8249 and 0.8509, respectively. The highest sensitivity was 0.070 6 °/(mg/dL), with a linear correlation coefficient of 0.993 and a low limit of detection of 0.2 mg/dL. Furthermore, binding affinity increased dramatically, with the Langmuir constants of 14.29 µM−1 for the gold-GQD probe and 0.000 1 µM−1 for the gold-APTES-GQD probe, representing a 142 900-fold increase. The probe demonstrated notable reproducibility and repeatability with relative standard deviations of 0.166% and 0.013%, respectively, and exceptional temporal stability of 99.66%. These findings represented a transformative leap in plasmonic UA sensors, characterized by enhanced precision, reliability, sensitivity, and increased surface binding capacity, synergistically fostering unprecedented practicality.

Sodium Enrichment of Mercury's Subsurface Through Diffusion

AbstractMercury's surface undergoes large temperature gradients between day and night, which repeats periodically over the same longitudes due to its 3:2 spin‐orbit resonance. This effect combined with the orbit's eccentricity, creates hot and cold geographic longitudes. The planet is covered with a highly porous regolith, allowing exospheric atoms to diffuse in depth. By using a 1‐D diffusion model, we studied the subsurface precipitation of gas over the cold and hot longitudes to understand gas retention. This work identifies the cold longitudes as favorable regions to form subsurface reservoirs closer to the surface. Moreover, subsurface reservoirs of adsorbates increase two to three times faster over cold longitudes than over hot longitudes, depending on the surface binding energy distribution of the atoms. We suggest that this result may be related to the observation that Mercury's sodium exosphere persists at later local times over the cold pole.

A mechanistic model of in vitro plasma activation to evaluate therapeutic kallikrein-kinin system inhibitors.

The kallikrein-kinin system (KKS) is a complex biochemical pathway that plays a crucial role in regulating several physiological processes, including inflammation, coagulation, and blood pressure. Dysregulation of the KKS has been associated with several pathological conditions such as hereditary angioedema (HAE), hypertension, and stroke. Developing an accurate quantitative model of the KKS may provide a better understanding of its role in health and disease and facilitate the rapid and targeted development of effective therapies for KKS-related disorders. Here, we present a novel, detailed mechanistic model of the plasma KKS, elucidating the processes of Factor XII (FXII) activation, the kallikrein feedback loop, cleavage of high molecular weight kininogen leading to bradykinin (BK) production, and the impact of inhibitors. The model incorporates both surface and solution-phase reactions of all proteins in the KKS, describing how binding site concentration affects the rate of surface reactions. The model was calibrated and validated using a variety of published and in-house experimental datasets, which encompass a range of dextran sulphate (DXS) concentrations to initiate contact activation and various KKS inhibitors to block bradykinin production. Our mathematical model showed that a trace amount of activated FXII is required for subsequent FXII activation. The model also reveals a bell-shaped curve relationship between the activation of the KKS and the number of DXS surface binding sites. Simulations of BK generation in healthy and HAE plasma demonstrated the impact of C1 esterase inhibitor (C1inh) deficiency via increased peak BK levels and accelerated formation in HAE plasma. The efficacy of KKS inhibitors, such as CSL312, ecallantide, and C1inh, was also evaluated, with CSL312 showing the most potent inhibition of BK generation. The present model represents a valuable framework for studying the intricate interactions within the plasma KKS and provides a better understanding of the mechanism of action of various KKS-targeted therapies.

S-palmitoylation of MAP kinase is essential for fungal virulence.

S-palmitoylation is an important reversible protein post-translational modification in organisms. However, its role in fungi is uncertain. Here, we found the treatment of the rice false fungus Ustilaginoidea virens with S-palmitoylation inhibitor 2 BP resulted in a significant decrease in fungal virulence. Comprehensive identification of S-palmitoylation sites and proteins in U. virens revealed a total of 4,089 S-palmitoylation sites identified among 2,192 proteins and that S-palmitoylated proteins were involved in diverse biological processes. Among the five palmitoyltransferases, UvPfa3 and UvPfa4 were found to regulate the pathogenicity of U. virens. We then performed quantitative proteomic analysis of ∆UvPfa3 and ∆UvPfa4 mutants. Interestingly, S-palmitoylated proteins were significantly enriched in the mitogen-activated protein kinase and autophagy pathways, and MAP kinase UvSlt2 was confirmed to be an S-palmitoylated protein which was palmitoylated by UvPfa4. Mutations of S-palmitoylation sites in UvSlt2 resulted in significantly reduced fungal virulence and decreased kinase enzymatic activity and phosphorylation levels. Simulations of molecular dynamics demonstrated mutation of S-palmitoylation sites in UvSlt2 causing decreased hydrophobic solvent-accessible surface area, thereby weakening the bonding force with its substrate UvRlm1. Taken together, S-palmitoylation promotes U. virens virulence through palmitoylation of MAP kinase UvSlt2 by palmitoyltransferase UvPfa4. This enhances the enzymatic phosphorylation activity of the kinase, thereby increasing hydrophobic solvent-accessible surface area and binding activity between the UvSlt2 enzyme and its substrate UvRlm1. Our studies provide a framework for dissecting the biological functions of S-palmitoylation and reveal an important role for S-palmitoylation in regulating the virulence of the pathogen.IMPORTANCES-palmitoylation is an important post-translational lipid modification of proteins. However, its role in fungi is uncertain. In this study, we found that S-palmitoylation promotes virulence of rice false smut fungus U. virens through palmitoylation of MAP kinase UvSlt2 by palmitoyltransferase UvPfa4. This enhances the enzymatic phosphorylation activity of the kinase, thereby increasing hydrophobic solvent-accessible surface area and binding activity between the UvSlt2 enzyme and its substrate UvRlm1. Our studies provide a framework for dissecting the biological functions of S-palmitoylation and reveal an important role for S-palmitoylation in regulating the virulence of the pathogen. This is the first functional study to reveal the role of S-palmitoylation in fungal virulence.

Comprehensive Screening of Plasma-Facing Materials for Nuclear Fusion

Plasma-facing materials (PFMs) represent one of the most significant challenges for the design of future nuclear fusion reactors. Inside the reactor, the divertor will experience the harshest material environment: intense bombardment of neutrons and plasma particles coupled with extremely large and intermittent heat fluxes. The material designated to cover this role in ITER is tungsten. While no other materials have shown the potential to match the properties of tungsten, many drawbacks associated with its application remain, including cracking and erosion induced by a low recrystallization temperature combined with a high ductile-brittle transition temperature and neutron-initiated embrittlement; surface morphology changes (helium bubbles and fuzz layer) due to plasma-tungsten interaction with subsequent risk of spontaneous material melting and delamination; and low oxidation resistance in the case of air contamination. Exploring alternatives to tungsten requires the design of a multivariable optimization problem. This work aims to produce a structured and comprehensive material screening of PFM candidates based on known inorganic materials. The method applied in this study to identify the most promising PFM candidates combines peer-reviewed data present in the PAULING FILE database and first-principles density-function theory calculations focusing on two key PFM defects—namely, the sputtering of surface atoms and the incorporation of interstitial hydrogen, respectively characterized by the surface binding energy and the interstitial formation energy. The crystal structures and their related properties, extracted from the PAULING FILE, are ranked according to the heat-balance equation of a PFM subjected to the heat loads in the divertor region of an ITER-like tokamak. The materials satisfying the requirements are critically compared with the state-of-the-art in plasma-facing materials research and in studies of refractory materials exposed to high temperatures, plasma, and neutron bombardment. This comparison assesses their thermo-mechanical properties under such conditions, identifying a subset of promising materials for first-principles electronic structure calculations. Most previously known and extensively studied PFMs, such as tungsten, molybdenum, and carbon-based materials, are captured by this screening process, confirming its reliability. Additionally, less familiar refractory materials suggest performance that calls for further investigations. Published by the American Physical Society 2024

Rational design of α-MnO2@TiO2 composites for high-performance zinc ions batteries

Aqueous zinc ions batteries (AZIBs) have attracted widespread interest due to their remarkable energy density, non-toxicity, environmental friendliness and low cost. However, the application of MnO2 cathode materials for AZIBs is limited owing to the dissolution of Mn2+, structural collapse and intrinsic poor conductivity. Herein, α-MnO2@TiO2 composites were synthesized via simple hydrothermal and calcination approaches. Some smaller TiO2 were coated on the MnO2 nanorods to form the protective layer. The TiO2 protective layer not only prevents the dissolution of Mn2+ and structural collapse of MnO2, but also enhances the conductivity of the material due to the presence of Ti3+ defects, which increased electrode materials wettability, specific surface area and diffusion rate of Zn2+. In addition, the coating TiO2 causes surface adsorbed oxygen and surface binding defects in α-MnO2@TiO2, and increases surface reactivity and electrochemically active sites, which provides abundant channels and available insertion electrochemically active sites for H+/Zn2+ transport. As a result, α-MnO2@TiO2–2 delivers a high specific capacity of 182.90 mAh g−1 at 1 A g−1 after 600 cycles and outstanding rate capacity of 127.91 mAh g−1 at 5 A g−1. This work provides an idea for exploring high-performance cathode materials for AZIBs.

A Novel Monoclonal Antibody Against a Modified Vaccinia Ankara (MVA) Envelope Protein as a Tool for MVA Virus Titration by Flow Cytometry.

Modified vaccinia Ankara (MVA) virus is a widely used vaccine platform, making accurate titration essential for vaccination studies. However, the current plaque forming unit (PFU) assay, the standard for MVA titration, is prone to observer bias and other limitations that affect accuracy and precision. To address these challenges, we developed a new flow cytometry-based quantification method using a highly specific monoclonal antibody (mAb) for the detection of MVA-infected cells, as a more accurate titration assay. Through previous work, we serendipitously identified three MVA-specific hybridoma antibody clones, which we characterized through ELISA, immunoblot, and flow cytometry, confirming their specificity for MVA. Sequencing confirmed that each antibody was monoclonal, and mass spectrometry results revealed that all mAbs target the MVA cell surface binding protein (CSBP, MVA105L). We next optimized the titration protocol using the most effective mAb, 33C7 by refining culture conditions and staining protocols to enhance sensitivity and minimize background. Our optimized method demonstrated superior sensitivity, reliability, and reduced processing time when compared with the traditional PFU assay, establishing it as a more accurate and efficient approach for MVA titration.

Electrochemical Analysis of Amyloid Plaques and ApoE4 with Chitosan-Coated Gold Nanostars for Alzheimer's Detection.

Monitoring the progression of Alzheimer's disease (AD) is crucial for mitigating dementia symptoms, alleviating pain, and improving mobility. Traditionally, AD biomarkers like amyloid plaques are predominantly identified in cerebrospinal fluid (CSF) due to their concentrated presence. However, detecting these markers in blood is hindered by the blood-brain barrier (BBB), resulting in lower concentrations. To address this challenge and identify pertinent AD biomarkers-specifically amyloid plaques and apolipoprotein E4 (ApoE4)-in blood plasma, we propose an innovative approach. This involves enhancing a screen-printed carbon electrode (SPCE) with an immobilization matrix comprising gold nanostars (AuNSs) coated with chitosan. Morphological and electrical analyses confirmed superior dispersion and conductivity with 0.5% chitosan, supported by UV-Vis spectroscopy, cyclic voltammetry, and Nyquist plots. Subsequent clinical assays measured electrical responses to quantify amyloid-β 42 (Aβ42) (15.63-1000 pg/mL) and APoE4 levels (0.41 to 40 ng/mL) in human blood plasma samples. Differential pulse voltammetry (DPV) responses exhibited peak currents proportional to biomarker concentrations, demonstrating high linear correlations (0.985 for Aβ42 and 0.919 for APoE4) with minimal error bars. Cross-reactivity tests with mixed solutions of amyloid-β 40 (Aβ40), Aβ42, and ApoE4 indicated minimal interference between biomarkers (<3% variation), further confirming the high specificity of the developed sensor. Validation studies demonstrated a strong concurrence with the gold-standard enzyme-linked immunosorbent assay (ELISA), while interference tests indicated a minimal variation in peak currents. This improved device presents promising potential as a point-of-care system, offering a less invasive, cost-effective, and simplified approach to detecting and tracking the progression of AD. The substantial surface binding area further supports the efficacy of our method, offering a promising avenue for advancing AD diagnostics.

Determining the carbon detection limit with magnetic sector dynamic secondary ion mass spectrometry (SIMS)

Dynamic SIMS is often used to evaluate the concentration of impurities in solids because of its high sensitivity, depth profiling capabilities, good depth resolution, and high throughput. Continuous ion beam sputtering with a high-density primary beam provides high sensitivity and reduces the background contribution from residual gases within the analytical chamber. Dynamic SIMS experiments performed with several magnetic sector instruments using various sputtering rate conditions demonstrated a slow decrease of the carbon detection limit with the increase of sputtering rate (SR), when it was varied by changing the sputtering beam density. The dependence data, within the scatter limits, can be fit by a power function of SR with the exponent of about −1/2. The observed detection-limit-dependence curve is common for magnetic sector SIMS instruments, owing to contamination and the design of the analytical chamber. The depth resolution of light-element SIMS analysis (except for oxygen) performed using Cs+ sputtering is limited and cannot be improved by reducing the impact energy of the sputtering beam. A segregation-type depth profile with a long trailing edge was observed in the B, C, and N depth profiles when Cs+ sputtering was employed under ultra-high vacuum conditions. This effect is plausibly associated with preferential sputtering owing to the difference in the surface binding energy, along with the large difference in the mass of the interacting atoms within the collision cascade during sputtering. The depth resolution improved with surface oxidation when Cs + sputtering was combined with backfilling of the analytical chamber using O2. The analytical chamber backfilled with oxygen can be sufficiently replaced with a very low-impact energy-focused oxygen beam, enabling simultaneous oxygen and cesium co-sputtering SIMS analysis. The practical feasibility of using a co-sputtering with focused Cs+ and O2+ sputtering beams to achieve a high depth-resolution for carbon analysis under ultrahigh vacuum conditions was demonstrated.

Theory of wetting dynamics with surface binding

Abstract Biomolecules, such as proteins and nucleic acids, can phase separate in the cytoplasm of cells to form biomolecular condensates. Such condensates are often liquid-like droplets that can wet biological surfaces such as membranes. Many molecules that participate in phase separation can also reversibly bind to membrane surfaces. When a droplet wets a surface, molecules can diffuse inside and outside of the droplet or in the bound state on the surface. How the interplay between surface binding, diffusion in surface and bulk affects the wetting kinetics is not well understood. Here, we derive the governing equations using non-equilibrium thermodynamics by relating the thermodynamic fluxes and forces at the surface coupled to the bulk. We study the spreading dynamics in the presence of surface binding and find that binding speeds up wetting by nucleating a droplet inside the surface. Our results suggest that the wetting dynamics of droplets can be regulated by two-dimensional surface droplets in the surface-bound layer through changing the binding affinity to the surfaces. These findings are relevant both to engineering life-like systems with condensates and vesicles, and biomolecular condensates in living cells.

An "R-spondin code" for multimodal signaling ON-OFF states.

R-spondins (RSPOs) are a family of secreted proteins and stem cell growth factors that are potent co-activators of Wnt signaling. Recently, RSPO2 and RSPO3 were shown to be multifunctional, not only amplifying Wnt- but also binding BMP- and FGF receptors to downregulate signaling. The common mechanism underlying these diverse functions is that RSPO2 and RSPO3 act as "endocytosers" that link transmembrane proteins to ZNRF3/RNF43 E3 ligases and trigger target internalization. Thus, RSPOs are natural protein targeting chimeras for cell surface proteins. Conducting data mining and cell surface binding assays we report additional candidate RSPO targets, including SMO, PTC1,2, LGI1, ROBO4, and PTPR(F/S). We propose that there is an "R-spondin code" that imparts combinatorial signaling ON-OFF states of multiple growth factors. This code involves the modular RSPO domains, notably distinct motifs in the divergent RSPO-TSP1 domains to mediate target interaction and internalization. The RSPO code offers a novel framework for the understanding how diverse signaling pathways may be coordinately regulated in development and disease.

Assessment of the Effects of ZnO and CuO Engineered Nanoparticles on Physicochemical Properties of Volcanic Ash Soil and Phosphorus Availability

The presence of engineered nanoparticles (ENPs) in soil systems can modify their properties and the availability of nutrients. This study evaluated the effect of 1% CuO or ZnO ENPs on the physicochemical properties and on the phosphorus (P) adsorption–desorption processes of a volcanic ash soil (Lautaro; LAU). The dynamics of P were conducted through kinetic and isotherm batch experiments. The results showed that LAU soil with 1% CuO or ZnO ENPs increased pHH2O (from 5.67 to 6.03 and 6.82, respectively), electrical conductivity (from 0.119 to 0.143 and 0.150 dS m−1, respectively), Zn availability (597.7 times higher for LAU with 1% ZnO ENPs in relation to soil without ENPs), and Cu availability (41.8 times higher for LAU with 1% CuO ENPs in relation to soil without ENPs). Moreover, the presence of ENPs decreased Brunauer, Emmett, and Teller specific surface area. The adsorption kinetic studies of P on LAU soil without and with 1% ENPs fitted well to the Elovich model (r2 ≥ 0.923), which indicated a chemiadsorption mechanism, whereas the adsorption isotherms were described by Langmuir–Freundlich model (r2 ≥ 0.939). The desorption percentage was LAU &gt; LAU + 1% CuO–ENPs &gt; LAU + 1% ZnO–ENPs, demonstrating an increased stability of the P–soil surface binding with 1% ENPs. Co–existing NO3−, SeO42−, and SO42− anions did not generate a steric hindrance between P and LAU soil binding. Finally, both ENPs could alter the quality of the soil due to changes in their physicochemical properties and decrease the availability of P in volcanic ash soils.

Effect of Cl2 Radical on Dry Development of Spin-Coated Metal Oxide Resist.

In this study, the effects of Cl2 radicals on dry development of spin-coated metal oxide resist (MOR) and changes in its surface binding states were investigated to verify the mechanism of dry development. Dry development characteristics of tin hydroxide (Tin OH), which is one of the MOR candidates for next generation lithography, were investigated as functions of process time and temperature using a Cl2 radicals source. Non-UV-exposed Tin OH film showed a linear etch rate (1.77 nm/min) from the initial thickness of ∼50 nm, while the UV-exposed film showed slower etch behavior (1.46 nm/min) in addition to the increase of film thickness for up to 3 min during the Cl2 radical dry development. UV-exposed photoresist (PR) contained more oxygen (Sn-O bonding) in the film due to the removal of butyl compounds from the clusters during the UV exposure process. Therefore, due to the lower reaction of chlorine radicals with Sn-O in the UV-exposed Tin OH than the other bindings, the non-UV-exposed PR was preferentially removed compared to the UV-exposed PR. As the temperature decreases, the overall etch rate decreases, but the difference in etch rate between exposed and unexposed Tin OH becomes larger. Finally, at a substrate temperature of -20 °C, the non-UV-exposed Tin OH with a thickness of 50 nm was completely removed, while ∼30 nm thick PR remained for UV-exposed Tin OH. Eventually, a negative tone development was possible with Cl2 radical plasma due to the difference in activation energy between the UV-exposed and non-UV-exposed films. It is believed that dry development using Cl2 radicals will be one of the most important process techniques for next-generation patterning to remove problems such as pattern leaning, line edge roughness, residue, etc., caused by wet development.

Discovery of a highly specific radiolabeled antibody targeting B-cell maturation antigen: Applications in PET imaging of multiple myeloma.

Multiple myeloma (MM) is characterized by the uncontrolled proliferation of monoclonal plasma cells (PC) in the bone marrow (BM). B-cell maturation antigen (BCMA) is predominantly expressed in malignant plasma cells, and associated with the proliferation, survival, and progression of various myeloma cells. Given these important roles, BCMA emerges as an ideal target antigen for MM therapy. However, effective stratification of patients who may benefit from targeted BCMA therapy and real-time monitoring the therapeutic efficacy poses significant clinical challenge. This study aims to develop a BCMA targeted diagnostic modality, and preliminarily explore its potential value in the radio-immunotherapy of MM. Using zirconium-89 (89Zr, t1/2 = 78.4 h) for labeling the BCMA-specific antibody, the BCMA-targeting PET tracer [89Zr]Zr-DFO-BCMAh230430 was prepared. The EC50 values of BCMAh230430 and DFO-BCMAh230430 were determined by ELISA assay. BCMA expression was assessed in four different tumor cell lines (MM.1S, RPMI 8226, BxPC-3, and KYSE520) through Western blot and flow cytometry. In vitro binding affinity was determined by cell uptake studies of [89Zr]Zr-DFO-BCMAh230430 in these tumor cell lines. For in vivo evaluation, PET imaging and ex vivo biodistribution studies were conducted in tumor-bearing mice to evaluate imaging performance and systemic distribution of [89Zr]Zr-DFO-BCMAh230430. Immunochemistry analysis was performed to detect BCMA expression in tumor tissues, confirming the specificity of our probe. Furthermore, we explored the anti-tumor efficacy of Lutetium-177 labeled BCMA antibody, [177Lu]Lu-DTPA-BCMAh230430, in tumor bearing-mice to validate its radioimmunotherapy potential. The radiolabeling of [89Zr]Zr-DFO-BCMAh230430 and [177Lu]Lu-DTPA-BCMAh230430 showed satisfactory radiocharacteristics, with a radiochemical purity exceeding 99%. ELISA assay results revealed closely aligned EC50 values for BCMAh230430 and DFO-BCMAh230430, which are 57 pM and 67 pM, respectively. Western blot and flow cytometry analyses confirmed the highest BCMA expression level. Cell uptake data indicated that MM.1S cells hada total cellular uptake (the sum of internalization and surface binding) of 38.3% ± 1.53% for [89Zr]Zr-DFO-BCMAh230430 at 12 h. PET imaging of [89Zr]Zr-DFO-BCMAh230430 displayed radioactive uptake of 7.71 ± 0.67%ID/g in MM.1S tumors and 4.13 ± 1.21%ID/g in KYSE520 tumors at 168 h post-injection (n = 4) (P < 0.05), consistent with ex vivo biodistribution studies. Immunohistochemical analysis of tumor tissues confirmed higher BCMA expression in MM.1S tumors xenograft compared to KYSE520 tumors. Notably, [177Lu]Lu-DTPA-BCMAh230430 showed some anti-tumor efficacy, evidenced by slowed tumor growth. Furthermore, no significant difference in body weight was observed in MM.1S tumor-bearing mice over 14 days of administration with or without [177Lu]Lu-DTPA-BCMAh230430. Our study has successfully validated the essential role of [89Zr]Zr-DFO-BCMAh230430 in non-invasively monitoring BCMA status in MM tumors, showing favorable tumor uptake and specific binding affinity to MM tumors. Furthermore, our research revealed, as a proof-of-concept, the effectiveness of [177Lu]Lu-DTPA-BCMAh230430 in radioimmunotherapy for MM tumors. In conclusion, we present a novel BCMA antibody-based radiotheranostic modality that holds promise for achieving efficient and precise MM diagnostic and therapy.

Simulation of ion beam sputtering with OKSANA and SRIM: A comparative study

The energy distributions and average energies of sputtered particles are calculated using simulation codes OKSANA and SRIM-2013. Most simulations refer to an amorphous Ni target bombarded by normally incident Ar ions of keV energies. Sputtering of Si, Ti, V and Nb targets is also considered. It is shown that SRIM can strongly overestimate the contribution of fast ejected atoms, especially for targets irradiated with lighter ions. The effect is amplified by using the surface binding energies found to fit the measured sputter yields. It is concluded that the enhanced ejection of fast particles is associated with sputtering due to backscattered ions. The role of the first collision of an incident ion with a target atom is particularly noted. A comparison of the OKSANA calculated results with the data of other codes (TRIM.SP, ACAT) and experimental data showed their reasonable agreement.

Functional Linkers Support Targeting of Multivalent Tweezers to Taspase1.

Taspase 1 is a unique protease not only pivotal for embryonic development but also implicated in leukemias and solid tumors. As such, this enzyme is a promising while still challenging therapeutic target, and with its protein structure featuring a flexible loop preceding the active site a versatile model system for drug development. Supramolecular ligands provide a promising complementary approach to traditional small-molecule inhibitors. Recently, the multivalent arrangement of molecular tweezers allowed the successful targeting of Taspase 1's surface loop. With this study we now want to take the next logic step und utilize functional linker systems that not only allow the implementation of novel properties but also engage in protein surface binding. Consequently, we chose two different linker types differing from the original divalent assembly: a backbone with aggregation-induced emission (AIE) properties to enable monitoring of binding and a calix[4]arene scaffold initially pre-positioning the supramolecular binding units. With a series of four AIE-equipped ligands with stepwise increased valency we demonstrated that the functionalized AIE linkers approach ligand binding affinities in the nanomolar range and allow efficient proteolytic inhibition of Taspase 1. Moreover, implementation of the calix[4]arene backbone further enhanced the ligands' inhibitory potential, pointing to a specific linker contribution.

The enigma of factor XII surface binding

The enigma of factor XII surface binding

Optimizing the method for expressing human monoclonal antibodies from a single peripheral blood cell from vaccinated donors

Human monoclonal antibodies are essential diagnostic and research tools and one of the most promising therapeutics. In the past years, single B cell technologies have evolved and over-come conventional methods' limitations, enabling the isolation of scarce B cell populations with desirable characteristics. In this study, we describe a simple and efficient method to isolate anti-gen-specific plasmablasts and memory B cells from hepatitis B virus vaccinated donors' peripheral blood and consequently amplification of immunoglobulin variable region genes. Amplified immunoglobulin variable region genes were cloned into expression vectors and transfected into a human cell line to produce human recombinant monoclonal antibodies. Major challenges in this protocol were isolation of antigen-specific B cells based on surface markers, recovering mRNA from a single cell for efficient amplification, and cloning the correct insert into a desired expression vector. The essential feature of our protocol was the separation of B cells from peripheral blood mononuclear cells before sorting. We identified antigen-specific binding B cells based on the expression of surface markers CD19, CD27, IgG, and binding to hepatitis B surface antigen. Efficient single-cell reverse transcription and polymerase chain reaction (RT-PCR) were achieved using a random primer mix and Kapa Hifi Hot Start Polymerase. Amplification efficiency differed among heavy and light chain variable regions (highest at heavy chain (68 %) and lowest at lambda light chain (22 %)). After co-transfection of HEK293T/17 with successfully cloned heavy and light chain vectors, 70 % of transfected cells produced recombinant monoclonal antibodies at a concentration ∼ 4 μg/ml and 7 % of them showed binding to HBsAg. Human monoclonal antibodies from peripheral blood have advantages over antibodies of mouse origin or phage display libraries, because of their high specificity and self-tolerance. Using the described protocol, we can generate fully human monoclonal antibodies to any other antigen for application in immunotherapy or basic research.

SC134-TCB Targeting Fucosyl-GM1, a T Cell-Engaging Antibody with Potent Antitumor Activity in Preclinical Small Cell Lung Cancer Models.

Small cell lung cancer (SCLC) is an aggressive disease with limited treatment options. Fucosyl-GM1 (FucGM1) is a glycolipid overexpressed in the majority of SCLC tumors but virtually absent from normal healthy tissues. In this study, we validate a FucGM1-targeting T cell-redirecting bispecific (TCB) antibody for the treatment of SCLC. More than 80% of patient-derived xenograft tissues of SCLC expressed FucGM1, whereas only three normal human tissues: pituitary, thymus, and skin expressed low and focal FucGM1. A FucGM1-targeting TCB (SC134-TCB), based on the Fc-silenced humanized SC134 antibody, exhibited nanomolar efficiency in FucGM1 glycolipid and SCLC cell surface binding. SC134-TCB showed potent ex vivo killing of SCLC cell lines with donor-dependent EC50 ranging from 7.2 pmol/L up to 211.0 pmol/L, effectively activating T cells, with picomolar efficiency, coinciding with target-dependent cytokine production such as IFNγ, IL2, and TNFα and robust proliferation of both CD4 and CD8 T cells. The ex vivo SC134-TCB tumor controlling activity translated into an effective in vivo anti-DMS79 tumor therapy, resulting in 100% tumor-free survival in a human peripheral blood mononuclear cell admixed setting and 40% overall survival (55% tumor growth inhibition) with systemically administered human peripheral blood mononuclear cells. Combination treatment with atezolizumab further enhanced survival and tumor growth inhibition (up to 73%). A 10-fold SC134-TCB dose reduction maintained the strong in vivo antitumor impact, translating into 70% overall survival (P < 0.0001). Whole-blood incubation with SC134-TCB, as well as healthy human primary cells analysis, revealed no target-independent cytokine production. SC134-TCB presents an attractive candidate to deliver an effective immunotherapy treatment option for patients with SCLC.

Cyclic Multi‐Site Chelation for Efficient and Stable Inverted Perovskite Solar Cells

Trap‐assisted non‐radiative recombination losses and moisture‐induced degradation significantly impede the development of highly efficient and stable inverted (p‐i‐n) perovskite solar cells (PSCs), which require high‐quality perovskite bulk. In this research, we mitigate these challenges by integrating thermally stable perovskite layers with Lewis base covalent organic frameworks (COFs). The ordered pore structure and surface binding groups of COFs facilitate cyclic, multi‐site chelation with undercoordinated lead ions, enhancing the perovskite quality across both its bulk and grain boundaries. This process not only reduces defects but also promotes improved energy alignment through n‐type doping at the surface. The inclusion of COF dopants in p‐i‐n devices achieves power conversion efficiencies (PCEs) of 25.64% (certified 24.94%) for a 0.0748‐cm2 device and 23.49% for a 1‐cm2 device. Remarkably, these devices retain 81% of their initial PCE after 978 hours of accelerated aging at 85˚C, demonstrating remarkable durability. Additionally, COF‐doped devices demonstrate excellent stability under illumination and in moist conditions, even without encapsulation.