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Polyacrylamide gel electrophoresis (PAGE) is a ubiquitous technique used in biochemical research laboratories to characterize protein samples. Despite its popularity, PAGE is relatively slow and provides limited separation resolution, especially for native proteins. This report describes the development of a microfluidic thermal gel transient isotachophoresis (TG-tITP) method to rapidly separate native proteins with high resolution. Thermal gels were employed as a separations matrix because of their unique ability to change viscosity in response to temperature. Proteins were added into thermal gel and loaded into a microfluidic device. Electrolyte optimization was conducted to achieve robust tITP to isotachophoretically preconcentrate proteins and then electrophoretically separate them. Electropherograms were collected through both time and distance to enable both small and large proteins to be measured within a single analysis. The effects of temperature were evaluated and found to exhibit a pronounced effect on the separation. Temperature gradients were then employed to alter thermal gel viscosity over time to maximize separation resolution between proteins. The results herein demonstrate how gradient TG-tITP achieves rapid, high-performance separations of native proteins. This analysis provided a wide mass range (6–464 kDa) with two-fold higher resolution than native PAGE while requiring 15,000-fold less protein loading and providing five-fold faster analysis times. Graphical abstract

In this study, a new hybrid hydrogel based on PAM (polyacrylamide)-Ag-g/WS[sub.2]/Ti[sub.3]C[sub.2]T[sub.x] was synthesized by radical polymerization using a conductive heterostructural nanocomposite WS[sub.2]/Ti[sub.3]C[sub.2]T[sub.x]. The synergy between the polymer matrix and the interface between two-dimensional nanomaterials ensured the production of a hydrogel with high extensibility and conductivity, as well as sensory characteristics. The composite hydrogel exhibited excellent strain-sensing capabilities, with gauge factors of 1.4 at low strain and 2.8 at higher strain levels. In addition, the material showed a fast response time of 2.17 s and a short recovery time of 0.46 s under cyclic stretching, which confirms its high reliability and reproducibility. The integration of Ti[sub.3]C[sub.2]T[sub.x] and WS[sub.2] promoted the formation of a conductive network in the hydrogel structure, which simultaneously increased its mechanical strength and signal stability under variable loads. Measurements confirm some potential of the PAM-Ag-g/WS[sub.2]/Ti[sub.3]C[sub.2]T[sub.x] composite hydrogel as a flexible wearable strain sensor. Based on measured numbers, we discussed the impact of the WS[sub.2]/Ti[sub.3]C[sub.2]T[sub.x] interface on the Gauge factor and conductivity of the composite. Theoretical modeling demonstrates significant changes in the electronic structure of the WS[sub.2]/Ti[sub.3]C[sub.2]T[sub.x] interface, and especially the WS[sub.2] surface, induced by substrate strain. Possible applications of the peculiar properties of PAM-Ag-g/WS[sub.2]/Ti[sub.3]C[sub.2]T[sub.x] composite were proposed.

The mitochondrial oxidative phosphorylation (OXPHOS) system plays a pivotal role in the cell’s energy conversion. The enzymes involved in OXPHOS are arranged in five protein-lipid complexes. The first four complexes (I–IV) form the mitochondrial respiratory chain, while Complex V is an F 1 F o -ATP synthase. Mutations in genes involved in the biosynthesis of the OXPHOS complexes are an important cause of metabolic diseases. Blue-native polyacrylamide gel electrophoresis (BN-PAGE), originally developed by Hermann Schägger in the 1990s, has become instrumental in gaining insights into structure/function relationships of the OXPHOS system, including: (1) the assembly pathways of the complexes, (2) the composition of higher-order respiratory chain supercomplexes and (3) pathologic mechanisms in patients with a monogenetic OXPHOS disorder. We have used BN-PAGE for >20 years and validate here our recently published step-by-step laboratory protocol. This protocol describes the manual casting of native mini-gels and sample preparation for the resolution of individual OXPHOS complexes or respiratory chain supercomplexes. In addition to BN-PAGE, we explain the closely related clear-native (CN)-PAGE and two-dimensional BN/denaturing-PAGE techniques. Downstream applications include western blot analysis and in-gel enzyme activity staining for Complexes I, II, IV and V. Limitations of the technique are the comparative insensitivity of in-gel Complex IV activity staining and the lack of in-gel Complex III activity staining. Compared to other published BN-PAGE protocols, our protocol contains a shortened sample extraction procedure, advises when to use BN-PAGE and when to use CN-PAGE, and suggests a simple enhancement step for in-gel Complex V activity staining that markedly improves sensitivity. Our protocol is adaptable and yields robust, semi-quantitative and reproducible results.

Polymers play a significant role in enhanced oil recovery (EOR) due to their viscoelastic properties and macromolecular structure. Herein, the mechanisms of the application of polymeric materials for enhanced oil recovery are elucidated. Subsequently, the polymer types used for EOR, namely synthetic polymers and natural polymers (biopolymers), and their properties are discussed. Moreover, the numerous applications for EOR such as polymer flooding, polymer foam flooding, alkali–polymer flooding, surfactant–polymer flooding, alkali–surfactant–polymer flooding, and polymeric nanofluid flooding are appraised and evaluated. Most of the polymers exhibit pseudoplastic behavior in the presence of shear forces. The biopolymers exhibit better salt tolerance and thermal stability but are susceptible to plugging and biodegradation. As for associative synthetic polyacrylamide, several complexities are involved in unlocking its full potential. Hence, hydrolyzed polyacrylamide remains the most coveted polymer for field application of polymer floods. Finally, alkali–surfactant–polymer flooding shows good efficiency at pilot and field scales, while a recently devised polymeric nanofluid shows good potential for field application of polymer flooding for EOR.

The objective of this study was to isolate and characterize collagen and angiotensin-I-converting enzyme (ACE)-inhibitory (ACEi) peptides from the swim bladders of monkfish (Lophius litulon). Therefore, acid-soluble collagen (ASC-M) and pepsin-soluble collagen (PSC-M) with yields of 4.27 ± 0.22% and 9.54 ± 0.51%, respectively, were extracted from monkfish swim bladders using acid and enzyme methods. The ASC-M and PSC-M contained Gly (325.2 and 314.9 residues/1000 residues, respectively) as the major amino acid, but they had low imino acid content (192.5 and 188.6 residues/1000 residues, respectively) in comparison with collagen from calf skins (CSC) (216.6 residues/1000 residues). The sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) patterns and ultraviolet (UV) absorption spectrums of ASC-M and PSC-M illustrated that they were mainly composed of type I collagen. Subsequently, three ACEi peptides were isolated from a PSC-M hydrolysate prepared via a double-enzyme system (alcalase + neutrase) and identified as SEGPK (MHP6), FDGPY (MHP7) and SPGPW (MHP9), with molecular weights of 516.5, 597.6 and 542.6 Da, respectively. SEGPK, FDGPY and SPGPW displayed remarkable anti-ACE activity, with IC50 values of 0.63, 0.94 and 0.71 mg/mL, respectively. Additionally, a molecular docking assay demonstrated that the affinities of SEGPK, FDGPY and SPGPW with ACE were −7.3, −10.9 and −9.4 kcal/mol, respectively. The remarkable ACEi activity of SEGPK, FDGPY and SPGPW was due to their connection with the active pockets and/or sites of ACE via hydrogen bonding, hydrophobic interaction and electrostatic force. Moreover, SEGPK, FDGPY and SPGPW could protect HUVECs by controlling levels of nitric oxide (NO) and endothelin-1 (ET-1). Therefore, this work provides an effective means for the preparation of collagens and novel ACEi peptides from monkfish swim bladders, and the prepared ACEi peptides, including SEGPK, FDGPY and SPGPW, could serve as natural functional components in the development of health care products to control hypertension.

The tarnished plant bug, Lygus lineolaris, and the red-banded stink bug, Piezodorus guildinii, pose significant economic threats to cotton and soybean crops in the mid-southern USA. However, the efficacy of insecticide spraying is comparatively low, and adjuvants play a crucial role in optimizing insecticide performance. This study evaluated the impact of two adjuvants, sodium alginate (SA) and polyacrylamide (PAM), on enhancing the efficacy of bifenthrin and imidacloprid via laboratory spray bioassays. Both SA and PAM demonstrated insignificant variation in LC[sub.50] values with formulated bifenthrin and imidacloprid. However, SA and PAM exhibited synergistic effects with two technical-grade insecticides. High concentrations of PAM increased the efficacy of bifenthrin by 1.50- and 1.70-fold for L. lineolaris and P. guildinii, respectively. Conversely, no enhancement effect was observed for the SA–technical-grade bifenthrin combination against either insect pests. Additionally, both SA and PAM enhanced the effectiveness of imidacloprid in P. guildinii by up to 2.68- and 2.73-fold, respectively. While a high concentration of PAM had a 1.45-fold synergistic effect on technical-grade imidacloprid, no enhancement effect was observed for the SA/imidacloprid combination in L. lineolaris. This study explored the synergistic impact of SA and PAM on the efficacy of technical-grade and formulated bifenthrin and imidacloprid, providing valuable insights into optimizing pest control strategies in agriculture.

Key messageA set of eight SNP markers was developed to facilitate the early selection of HMW-GS alleles in breeding programmes. In bread wheat (Triticum aestivum), the high molecular weight glutenin subunits (HMW-GSs) are the most important determinants of technological quality. Known to be very diverse, HMW-GSs are encoded by the tightly linked genes Glu-1-1 and Glu-1-2. Alleles that improve the quality of dough have been identified. Up to now, sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS-PAGE) of grain proteins is the most widely used for their identification. To facilitate the early selection of HMW-GS alleles in breeding programmes, we developed DNA-based molecular markers. For each accession of a core collection (n = 364 lines) representative of worldwide bread wheat diversity, HMW-GSs were characterized by both genotyping and SDS-PAGE. Based on electrophoresis, we observed at least 8, 22 and 9 different alleles at the Glu-A1, Glu-B1 and Glu-D1 loci, respectively, including new variants. We designed a set of 17 single-nucleotide polymorphism (SNP) markers that were representative of the most frequent SDS-PAGE alleles at each locus. At Glu-A1 and Glu-D1, two and three marker-based haplotypes, respectively, captured the diversity of the SDS-PAGE alleles rather well. Discrepancies were found mainly for the Glu-B1 locus. However, statistical tests revealed that two markers at each Glu-B1 gene and their corresponding haplotypes were more significantly associated with the rheological properties of the dough than were the relevant SDS-PAGE alleles. To conclude, this study demonstrates that the SNP markers developed provide additional information on HMW-GS diversity. Two markers at Glu-A1, four at Glu-B1 and two at Glu-D1 constitute a useful toolbox for breeding wheat to improve end-use value.

Background: The present study aimed to fabricate surface-modified chitosan nanoparticles with two mucoadhesive polymers (sodium alginate and polyethylene glycol) to optimize their protein encapsulation efficiency, improve their mucoadhesion properties, and increase their stability in biological fluids. Method: Ionotropic gelation was employed to formulate chitosan nanoparticles and surface modification was performed at five different concentrations (0.05, 0.1, 0.2, 0.3, 0.4% w/v) of sodium alginate (ALG) and polyethylene glycol (PEG), with ovalbumin (OVA) used as a model protein antigen. The functional characteristics were examined by dynamic light scattering (DLS), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), and scanning electron microscopy (SEM)/scanning transmission electron microscopy (STEM). Stability was examined in the presence of simulated gastric and intestinal fluids, while mucoadhesive properties were evaluated by in vitro mucin binding and ex vivo adhesion on pig oral mucosa tissue. The impact of the formulation and dissolution process on the OVA structure was investigated by sodium dodecyl-polyacrylamide gel electrophoresis (SDS-PAGE) and circular dichroism (CD). Results: The nanoparticles showed a uniform spherical morphology with a maximum protein encapsulation efficiency of 81%, size after OVA loading of between 200 and 400 nm and zeta potential from 10 to 29 mV. An in vitro drug release study suggested successful nanoparticle surface modification by ALG and PEG, showing gastric fluid stability (4 h) and a 96 h sustained OVA release in intestinal fluid, with the nanoparticles maintaining their conformational stability (SDS-PAGE and CD analyses) after release in the intestinal fluid. An in vitro mucin binding study indicated a significant increase in mucin binding from 41 to 63% in ALG-modified nanoparticles and a 27–49% increase in PEG-modified nanoparticles. The ex vivo mucoadhesion showed that the powdered particles adhered to the pig oral mucosa. Conclusion: The ALG and PEG surface modification of chitosan nanoparticles improved the particle stability in both simulated gastric and intestinal fluids and improved the mucoadhesive properties, therefore constituting a potential nanocarrier platform for mucosal protein vaccine delivery.

This study investigates the interaction of polyacrylamide (PAM) of different functional groups (sulfonate vs. carboxylate) and charge density (30% hydrolysed vs. 10% hydrolysed) with calcium carbonate (CaCO3) via atomic force microscopy (AFM) and partly via molecular dynamic (MD) simulations. The PAM used were F3330 (30% hydrolysed), AN125 (25% sulfonated), and AN910 (% hydrolysed). A total of 100 ppm of PAMs was prepared in 0.1% NaCl, 3% NaCl, and 4.36% NaNO3 to be employed in AFM experiments, while oligomeric models (30 repeating units) of hydrolysed polyacrylamide (HPAM), sulfonated polyacrylamide (SPAM), and neutral PAM (NPAM) were studied on a model calcite surface on MD simulations. AFM analysis indicated that F3330 has a higher average adhesion and interaction energy with CaCO3 than AN125 due to the bulky sulfonate side group of AN125 interfering with SPAM adsorption. Steric repulsion of both PAMs was similar due to their comparable molecular weights and densities of the charged group. In contrast, AN910 showed lower average adhesion and interaction energy, along with slightly longer steric repulsion with calcite than F3330, suggesting AN910 adopts more loops and tails than the slightly flatter F3330 configuration. An increase in salt concentration from 0.1% to 3% NaCl saw a reduction in adhesion and interaction energy for F3330 and AN125 due to charge screening, while AN910 saw an increase, and these values increased further at 4.36% NaNO3. MD simulations revealed that the salt ions in the system formed salt bridges between PAM and calcite, indicating that the adhesion and interaction energy observed from AFM are likely to be the net balance between PAM charged group screening and salt bridging by the salt ions present. Salt ions with larger bare radii and smaller hydrated radii were shown to form stronger salt bridges.

Background The consumption of oats has rapidly increased due to their exceptional nutritional value. However, concerns over genetic erosion have emerged as oat breeding programs rely on a highly limited genetic pool. This study aimed to expand the genetic diversity pool of oats by collecting wild oat ( Avena fatua L.) populations in South Korea and assessing their genetic diversity and seed storage protein patterns. Results A total of 237 A. fatua individuals were collected in 2022 from eight regions in the southwestern coastal areas of South Korea. Genetic diversity and seed storage protein patterns were analyzed using genotyping-by-sequencing (GBS) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The GBS analysis identified 20,836 single-nucleotide polymorphisms (SNPs). An analysis of molecular variance (AMOVA) based on regional populations revealed that 40.9% of the genetic variation was attributed to differences among populations, while 59.1% was within populations, indicating high genetic differentiation within regional populations. Subsequent population structure analysis and discriminant analysis of principal components (DAPC) both stated the formation of two distinct genetic groups, with an AMOVA value of 70.9% between the groups, suggesting a high level of genetic variation. Pairwise F ST analysis was conducted to compare the genetic differentiation between two populations, revealing that Jindo and Jangheung exhibited the highest level of genetic differentiation ( F ST = 0.795) among the geographic groups. Seed storage proteins were analyzed using SDS-PAGE, and the patterns were grouped using k-means clustering. A comparison between the groups based on protein patterns and those based on genetic variation revealed no significant correlation. Conclusion This study provides data on the genetic diversity of A. fatua , a wild relative of cultivated oats, aimed at expanding the genetic pool of oats for future breeding programs. These findings are expected to be a foundational resource for oat breeding and genetic improvement efforts.