Microbead-Encapsulated Luminescent Bioreporter Screening of P. aeruginosa via Its Secreted Quorum-Sensing Molecules.

Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium that remains a prevalent clinical and environmental challenge. Quorum-sensing (QS) molecules are effective biomarkers in pinpointing the presence of P. aeruginosa. This study aimed to develop a convenient-to-use, whole-cell biosensor using P. aeruginosa reporters individually encapsulated within alginate-poly-L-lysine (alginate-PLL) microbeads to specifically detect the presence of bacterial autoinducers. The PLL-reinforced microbeads were prepared using a two-step method involving ionic cross-linking and subsequent coating with thin layers of PLL. The alginate-PLL beads showed good stability in the presence of a known cation scavenger (sodium citrate), which typically limits the widespread applications of calcium alginate. In media containing synthetic autoinducers-such as N-(3-oxo dodecanoyl) homoserine lactone (3-oxo-C12-HSL) and N-butanoyl-L-homoserine lactone (C4-HSL), or the cell-free supernatants of planktonic or the flow-cell biofilm effluent of wild P. aeruginosa (PAO1)-the encapsulated bacteria enabled a dose-dependent detection of the presence of these QS molecules. The prepared bioreporter beads remained stable during prolonged storage at 4 and -80 °C and were ready for on-the-spot sensing without the need for recovery. The proof-of-concept, optical fiber-based, and whole-cell biosensor developed here demonstrates the practicality of the encapsulated bioreporter for bacterial detection based on specific QS molecules.

Development of cashew-alginate microbeads and powdered dose forms: prospects for oral vaccine delivery in chickens.

Conventional oral vaccine delivery in poultry is challenging due to vaccine degradation in the gastrointestinal (GI) environment and the need for cold-chain storage. Microencapsulation offers a solution by protecting vaccines from GI degradation and improving stability. Natural polymers like alginate and cashew gum have mucoadhesive properties, making them promising candidates for oral vaccine delivery. This study developed cashew-alginate microbeads and a powdered dose form for oral vaccine delivery in chickens. The microbeads were created using ionotropic gelation, while the powdered form was obtained via freeze-drying. These formulations were characterized for size, shape, and stability using scanning electron microscopy (SEM), light microscopy, X-ray diffraction (XRD), and Energy Dispersive X-ray (EDX). Peak adhesion time (PAT) was determined using chicken intestinal and esophageal tissues, and antigenicity was assessed with in-vitro hemagglutination (HA) and hemagglutination inhibition (HI) assays. The microbeads exhibited a spherical shape with a porous structure, suggesting enhanced antigen accommodation. Hemagglutination Inhibition tests indicated that the experimental vaccine remained effective without cold-chain storage for three months. These findings suggest that cashew-alginate microbeads are promising for oral vaccine delivery in poultry.

Spatially Confined Assembly and Immobilization of Hierarchical Nanoparticle Architectures inside Microdroplets in Magnetic Fields.

Magnetic field-directed colloidal interactions offer facile tools for assembly of structures that range from linear chains to multidimensional hierarchical architectures. While the field-driven assembly of colloidal particles has commonly been investigated in unbounded media, a knowledge gap remains concerning such assembly in confined microenvironments. Here, we investigate how confinement of ferromagnetic nanoparticles in microspheres directs their magnetic assembly into hierarchical architectures. Microdroplets from polydimethylsiloxane (PDMS) liquid precursor containing dispersed iron oxide magnetic nanoparticles (MNPs) were placed in a static magnetic field leading to the formation of organized assemblies inside the host droplets. By changing the MNP concentrations, we revealed a sequence of microstructures inside the droplets, ranging from linear chains at a low MNP loading, transitioning to a combination of chains and networked bundles, to solely 3D bundles at high MNP loading. These experimental results were analyzed with the aid of COMSOL simulations where we calculated the potential energy to identify the preferred assembly conformations. The chains at high MNP loading minimized their energy by aggregating laterally to form bundles with their MNP dipoles being out-of-registry. We cured these PDMS droplets to immobilize the assemblies by forming soft microbeads. These microbeads constitute an "interaction toolbox" with different magnetic macroscale responses, which are governed by the structuring of the MNPs and their magnetic polarizability. We show that thanks to their ability to rotate by field-induced torque under a rotating field, these microbeads can be employed in applications such as optical modulators and microrollers.

Electrogenerated chemiluminescence from luminol-labelled microbeads triggered by in situ generation of hydrogen peroxide.

We developed a sensing strategy that mimics the bead-based electrogenerated chemiluminescence immunoassay. However, instead of the most common metal complexes, such as Ru or Ir, the luminophore is luminol. The electrogenerated chemiluminescence of luminol was promoted by in situ electrochemical generation of hydrogen peroxide at a boron-doped diamond electrode. The electrochemical production of hydrogen peroxide was achieved in a carbonate solution by an oxidation reaction, while at the same time, microbeads labelled with luminol were deposited on the electrode surface. For the first time, we proved that was possible to obtain light emission from luminol without its direct oxidation at the electrode. This new emission mechanism is obtained at higher potentials than the usual luminol electrogenerated chemiluminescence at 0.3-0.5 V, in conjunction with hydrogen peroxide production on boron-doped diamond at around 2-2.5 V (vs Ag/AgCl).

Betulin Stimulates Osteogenic Differentiation of Human Osteoblasts-Loaded Alginate-Gelatin Microbeads.

Osteoporosis, a terminal illness, has emerged as a global public health problem in recent years. The long-term use of bone anabolic drugs to treat osteoporosis causes multi-morbidity in elderly patients. Alternative therapies, such as allogenic and autogenic tissue grafts, face important issues, such as a limited source of allogenic grafts and tissue rejection in autogenic grafts. However, stem cell therapy has been shown to increase bone regeneration and decrease osteoporotic bone formation. Stem cell therapy combined with betulin (BET) supplementation might be adequate for bone remodeling and new bone tissue generation. In this study, the effect of BET on the viability and osteogenic differentiation of hFOB 1.19 cells was investigated. The cells were encapsulated in alginate-gelatin (AlGel) microbeads. In vitro tests were conducted during the 12 d of incubation. While BET showed cytotoxic activity (>1 µM) toward non-encapsulated hFOB 1.19 cells, encapsulated cells retained their functionality for up to 12 days, even at 5 µM BET. Moreover, the expression of osteogenic markers indicates an enhanced osteo-inductive effect of betulin on encapsulated hFOB 1.19, compared to the non-encapsulated cell culture. The 3D micro-environment of the AlGel microcapsules successfully protects the hFOB 1.19 cells against BET cytotoxicity, allowing BET to improve the mineralization and differentiation of osteoblast cells.

Stability and release mechanism of double emulsification (W1/O/W2) for biodegradable pH-responsive polyhydroxybutyrate/cellulose acetate phthalate microbeads loaded with the water-soluble bioactive compound niacinamide.

Microbeads of biodegradable polyhydroxybutyrate (PHB) offer environmental benefits and economic competitiveness. The aim of this study was to encapsulate a water-soluble bioactive compound, niacinamide (NIA), in a pH-responsive natural matrix composed of PHB and cellulose acetate phthalate (CAP) by double emulsification (W1/O/W2) to improve the encapsulation efficiency (%EE) and loading capacity (%LC). PHB was produced in-house by Escherichia coli JM109 pUC19-23119phaCABA-04 without the inducing agent isopropyl ß-D-1-thiogalactopyranoside (IPTG). The influences of PHB and polyvinyl alcohol (PVA) concentrations, stirring rate, PHB/CAP ratio and initial NIA concentration on the properties of NIA-loaded pH-responsive microbeads were studied. The NIA-loaded pH-responsive PHB/CAP microbeads exhibited a spherical core-shell structure. The average size of the NIA-loaded pH-responsive microbeads was 1243.3 ± 11.5 µm. The EE and LC were 33.3 ± 0.5 % and 28.5 ± 0.4 %, respectively. The release profiles of NIA showed pH-responsive properties, as 94.2 ± 3.5 % of NIA was released at pH 5.5, whereas 99.3 ± 2.4 % of NIA was released at pH 7.0. The NIA-loaded pH-responsive PHB/CAP microbeads were stable for >90 days at 4 °C under darkness, with NIA remaining at 73.65 ± 1.86 %. A cytotoxicity assay in PSVK1 cells confirmed that the NIA-loaded pH-responsive PHB/CAP microbeads were nontoxic at concentrations lower than 31.3 µg/mL, in accordance with ISO 10993-5.

Structural Color Inks Containing Photonic Microbeads for Direct Writing.

Printing structurally colored patterns is of great importance for providing customized graphics for various purposes. Although a direct writing technique has been developed, the use of colloidal dispersions as photonic inks requires delicate printing conditions and restricts the mechanical and optical properties of printed patterns. In this work, we produce elastic photonic microbeads through scalable bulk emulsification and formulate photonic inks containing microbeads for direct writing. To produce the microbeads, a photocurable colloidal dispersion is emulsified into a highly concentrated sucrose solution via vortexing, which results in spherical emulsion droplets with a relatively narrow size distribution. The microbeads are produced by photopolymerization and are then suspended in urethane acrylate resin at volume fractions of 0.35-0.45. The photonic inks retain high color saturation of the microbeads and offer enhanced printability and dimensional control on various target substrates including fabrics, papers, and even skins. Importantly, the printed graphics show high mechanical stability as the elastic microbeads are embedded in the polyurethane matrix. Moreover, the colors show a wide viewing angle and low-angle dependency due to the optical isotropy of individual microbeads and light refraction at the air-matrix interface. We postulate that this versatile direct writing technique is potentially useful for structural color coating and printing on the surfaces of arbitrary 3D objects.

Synthesis of eco-friendly multifunctional dextran microbeads for multiplexed assays.

There has been an increasing demand for simultaneous detection of multiple analytes in one sample. Microbead-based platforms have been developed for multiplexed assays. However, most of the microbeads are made of non-biodegradable synthetic polymers, leading to environmental and human health concerns. In this study, we developed an environmentally friendly dextran microbeads as a new type of multi-analyte assay platform. Biodegradable dextran was utilized as the primary material. Highly uniform magnetic dextran microspheres were successfully synthesized using the Shirasu porous glass (SPG) membrane emulsification technique. To enhance the amount of surface functional groups for ligand conjugation, we coated the dextran microbeads with a layer of dendrimers via a simple electrostatic adsorption process. Subsequently, a unique and efficient click chemistry coupling technique was developed for the fluorescence encoding of the microspheres, enabling multiplexed detection. The dextran microbeads were tested for 3-plex cytokine analysis, and exhibited excellent biocompatibility, stable coding signals, low background noise and high sensitivity.

Harnessing the Therapeutic Potential of Dillenia indica: An Overview of Recent Dosage Form Developments.

Dillenia indica, commonly known as Elephant Apple, is a significant medicinal plant found in Assam, North-East India. This evergreen shrub or small to medium-sized tree possesses not only tasty components but also a plethora of beneficial therapeutic characteristics. This review article aims to explore the potential use of Dillenia indica in the treatment of diabetes and other diseases, as well as discuss various patents associated with this plant. The study focuses on identifying different formulations derived from various parts of Dillenia indica. These formulations encompass a range of dosage forms, including mucoadhesive buccal dosage forms, buccal patches, microbeads, emulgel, and mucoadhesive nasal gel. Each of these dosage forms offers unique advantages and applications. Mucoadhesive buccal dosage forms are designed to adhere to the oral mucosa, allowing for controlled drug release and enhanced absorption. Buccal patches provide a convenient and localized delivery system for specific therapeutic agents. Microbeads offer a versatile approach for encapsulating drugs and facilitating their controlled release. Emulgels combine the benefits of both emulsions and gels, providing improved drug delivery and stability. Mucoadhesive nasal gels offer a non-invasive route for drug administration, allowing for rapid absorption through the nasal mucosa. By exploring these different formulations, researchers aim to harness the therapeutic potential of Dillenia indica in a variety of diseases, including diabetes. The study also highlights the importance of patents associated with Dillenia indica, indicating the growing interest in its medicinal properties and potential commercial applications. Dillenia indica holds promise as a valuable medicinal plant, with its diverse therapeutic characteristics and tasty components. The study discussed various formulations derived from different parts of the plant, showcasing their potential applications in the treatment of diseases. Further research and development in this field may lead to the discovery of novel treatments and contribute to the advancement of pharmaceutical science.

Microplastics in personal care products and cosmetics in Sri Lanka.

In the Sri Lankan context, the lack of baseline studies to mitigate microplastic emissions through personal care and cosmetic products poses a huge problem. Hence this study serves as the first scientific investigation to analyze and characterize microplastics in selected personal care and cosmetic items available in the Sri Lankan markets. Fifteen brands representing five categories (face wash, facial scrubs, baby creams, shaving creams, and skin creams) of personal care and cosmetic items served as the basis for this investigation. Based on a questionnaire survey, from each category, three highly utilized brands were chosen and triplicates from each brand were used for the analysis. All samples were treated with the Fenton reagent to extract microplastics. Then through Nile red staining suspected microplastic were screened and characterized through FT-IR spectroscopy. The Nile Red analysis revealed seven brands of the fifteen to be stained with Nile Red and demonstrate luminance properties under UV light. However, FT-IR analysis proved only six brands contained actual microplastics. Low-density polyethylene and ethylene-propylene copolymer were the dominant types of microplastic. Most microplastics were irregularly shaped and white in color with sizes ranging from 238.55 ± 50.74 to 450.69 ± 174.9 µm. An emission estimation revealed that products FS-01 and FW-03 contain 3.36 ± 0.20 g and 0.2 ± 0.05 g of isolatable microplastics per product. While the present study provides scientific evidence for the availability of microplastics in products in Sri Lankan markets, it also provides a great opportunity to develop relevant policies and regulations to control them.

Alginate- and Chitosan-Modified Gelatin Hydrogel Microbeads for Delivery of E. coli Phages.

Bacterial infections are among the most significant health problems/concerns worldwide. A very critical concern is the rapidly increasing number of antibiotic-resistant bacteria, which requires much more effective countermeasures. As nature's antibacterial entities, bacteriophages shortly ("phages") are very important alternatives to antibiotics, having many superior features compared with antibiotics. The development of phage-carrying controlled-release formulations is still challenging due to the need to protect their activities in preparation, storage, and use, as well as the need to create more user-friendly forms by considering their application area/site/conditions. Here, we prepared gelatin hydrogel microbeads by a two-step process. Sodium alginate was included for modification within the initial recipes, and these composite microbeads were further coated with chitosan. Their swelling ratio, average diameters, and Zeta potentials were determined, and degradations in HCl were demonstrated. The target bacteria Escherichia coli (E.coli) and its specific phage (T4) were obtained from bacterial culture collections and propagated. Phages were loaded within the microbeads with a simple method. The phage release characteristics were investigated comparatively and were demonstrated here. High release rates were observed from the gelatin microbeads. It was possible to reduce the phage release rate using sodium alginate in the recipe and chitosan coating. Using these gelatin-based microbeads as phage carrier matrices-especially in lyophilized forms-significantly improved the phage stability even at room temperature. It was concluded that phage release from gelatin hydrogel microbeads could be further controlled by alginate and chitosan modifications and that user-friendly lyophilized phage formulations with a much longer shelf life could be produced.

Nanocomposite microbeads made of recycled polylactic acid for the magnetic solid phase extraction of xenobiotics from human urine.

Nanocomposite microbeads (average diameter = 10-100 µm) were prepared by a microemulsion-solidification method and applied to the magnetic solid-phase extraction (m-SPE) of fourteen analytes, among pesticides, drugs, and hormones, from human urine samples. The microbeads, perfectly spherical in shape to maximize the surface contact with the analytes, were composed of magnetic nanoparticles dispersed in a polylactic acid (PLA) solid bulk, decorated with multi-walled carbon nanotubes (mPLA@MWCNTs). In particular, PLA was recovered from filters of smoked electronic cigarettes after an adequate cleaning protocol. A complete morphological characterization of the microbeads was performed via Fourier-transform infrared (FTIR) spectroscopy, UV-Vis spectroscopy, thermogravimetric and differential scanning calorimetry analysis (TGA and DSC), scanning electron microscopy (SEM) and X-ray diffraction analysis (XRD). The recovery study of the m-SPE procedure showed yields ≥ 64%, with the exception of 4-chloro-2-methylphenol (57%) at the lowest spike level (3 µg L-1). The method was validated according to the main FDA guidelines for the validation of bioanalytical methods. Using liquid chromatography-tandem mass spectrometry, precision and accuracy were below 11% and 15%, respectively, and detection limits of 0.1-1.8 µg L-1. Linearity was studied in the range of interest 1-15 µg L-1 with determination coefficients greater than 0.99. In light of the obtained results, the nanocomposite microbeads have proved to be a valid and sustainable alternative to traditional sorbents, offering good analytical standards and being synthetized from recycled plastic material. One of the main objectives of the current work is to provide an innovative and optimized procedure for the recycling of a plastic waste, to obtain a regular and reliable microstructure, whose application is here presented in the field of analytical chemistry. The simplicity and greenness of the method endows the procedure with a versatile applicability in different research and industrial fields.

Aptamer decorated PDA@magnetic silica microparticles for bacteria purification.

One significant constraint in the advancement of biosensors is the signal-to-noise ratio, which is adversely affected by the presence of interfering factors such as blood in the sample matrix. In the present investigation, a specific aptamer binding was chosen for its affinity, while exhibiting no binding affinity towards non-target bacterial cells. This selective binding property was leveraged to facilitate the production of magnetic microparticles decorated with aptamers. A novel assay was developed to effectively isolate S. pneumoniae from PBS or directly from blood samples using an aptamer with an affinity constant of 72.8 nM. The capture experiments demonstrated efficiencies up to 87% and 66% are achievable for isolating spiked S. pneumoniae in 1 mL PBS and blood samples, respectively.

Process Study of Selective Laser Sintering of PS/GF/HGM Composites.

To address the issues of insufficient strength and poor precision in polystyrene forming parts during the selective laser sintering process, a ternary composite of polystyrene/glass fiber/hollow glass microbeads was prepared through co-modification by incorporating glass fiber and hollow glass microbeads into polystyrene using a mechanical mixing method. The bending strength and dimensional accuracy of the sintered composites were investigated by conducting an orthogonal test and analysis of variance to study the effects of laser power, scanning speed, scanning spacing, and delamination thickness. The process parameters were optimized and selected to determine the optimal combination. The results demonstrated that when considering bending strength and Z-dimensional accuracy as evaluation criteria for terpolymer sintered parts, the optimum process parameters are as follows: laser power of 24 W, scanning speed of 1600 mm/s, scanning spacing of 0.24 mm, and delamination thickness of 0.22 mm. Under these optimal process parameters, the bending strength of sintered parts reaches 6.12 MPa with a relative error in the Z-dimension of only 0.87%. The bending strength of pure polystyrene sintered parts is enhanced by 15.69% under the same conditions, while the relative error in the Z-dimension is reduced by 63.45%. It improves the forming strength and precision of polystyrene in the selective laser sintering process and achieves the effect of enhancement and modification, which provides a reference and a new direction for exploring polystyrene-based high-performance composites and expands the application scope of selective laser sintering technology.

A dataset of microstructure features of electro-hydrodynamic assisted 5-fluorouracil-grafted alginate microbeads and physicochemical properties for effective colon targeted carriers drug delivery.

5-Fluorouracil (5-FU) has been the primary drug used in chemotherapy for colorectal carcinoma, and localizing the drug would be effective in avoiding its side effects and improving therapeutic outcomes. One approach to achieve this is by encapsulating the drug in microbeads. Alginate microbeads, in particular, exhibit promising pH-sensitive properties, making them an attractive option for colon targeting. Thus, the main aim of this study is to formulate and characterize 5-FU-encapsulated alginate microbeads as a pH-sensitive drug delivery system for controlled release in the gastrointestinal tract. In this study, the alginate microbeads encapsulating 5-FU was manufactured using electrospray methods. This method offers the advantages of promoting the formulation of uniformly small-sized microbeads with improved performance in terms of swelling and diffusion rates. The size and shape of the 5-FU microbeads are 394.23 ± 3.077 µm and have a spherical factor of 0.026 ± 0.022, respectively, which are considered acceptable and indicative of a spherical shape. The microbeads' encapsulation efficiency was found to be 69.65 ± 0.18%, which is considered high in comparison to other literature. The attenuated total reflectance - Fourier transform infrared spectroscopy (ATR-FTIR) data confirmed the complexation of sodium alginate with calcium ions, along with the encapsulation of 5-FU in the microbeads matrix. The 5-FU microbeads displayed pH-dependent swelling, exhibiting less swelling in simulated gastric fluid (SGF) than in simulated intestinal fluid (SIF). Additionally, the release of 5-FU from the microbeads is pH-dependent, with the cumulative percentage drug release being higher in simulated intestinal fluid than in SGF. The data indicate that the 5-FU microbeads can be utilized for the delivery of 5-FU in colon-targeted therapy, potentially leading to improved tumor treatment.

Dual-Encoded Affinity Microbead Signature Combinatorial Profiling for Acute Myocardial Infarction High-Sensitivity Diagnosis.

The early diagnosis of acute myocardial infarction (AMI) is dependent on the combined feedback of multiple cardiac biomarkers. However, it remains challenging to precisely detect multicardiac biomarkers in complex blood early due to the lack of sensitive and specific diagnostic indicators and the low abundance and small size of associated biomarkers with high specificity (such as microRNAs). To make matters worse, spectral overlap significantly limits the multiplex analysis of cardiac biomarkers by fluorescent probes, leading to bias in the diagnosis of myocardial infarction. Herein, we developed a method for simultaneous detection of miRNAs and protein biomarkers using size- and color-coded microbeads that carry signature for target capture. We also constructed a microfluidic chip with different spacer arrays that segregate these microbeads in different chip regions according to their size to produce signature signals, indicating the level of different biomarkers. The signals on the microbeads were hugely amplified by catalytic hairpin assembly and rolling circle amplification. Notably, this strategy enables the simultaneous and in situ sensitive profiling of six kinds of biomarkers via adding two different fluorescent labels, removing the limitations of spectral overlap. We envision that the strategy has great potential for application in clinical diagnosis for AMI.

Conception of pH-sensitive calcium alginate/poly vinyl alcohol hydrogel beads for controlled oral curcumin delivery systems. Antibacterial and antioxidant properties.

Curcumin, a bioactive compound derived from the rhizome of Curcuma longa, has gained widespread attention for its potential therapeutic properties, including anti-inflammatory, antioxidant and anticancer effects. However, its poor aqueous solubility, instability and limited bioavailability have hindered its clinical applications. New beads formulations based on sodium alginate biopolymer (SA) and poly vinyl alcohol (PVA) were successfully prepared and evaluated as a potential drug vehicle for extended release of curcumin (Cur). Pristine and curcumin loaded calcium alginate/poly vinyl alcohol beads (CA/PVA and CA/PVA/Cur) at different compositions of SA and PVA were prepared by an ionotropic gelation method of SA followed by two freeze-thawing (FT) cycles for further crosslinking of PVA. Characterization techniques, such as scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), UV-Visible spectroscopy, thermogravimetric analysis (TGA) and x-ray diffraction (XRD) were used to confirm the successful microencapsulation of curcumin within the CA/PVA microcapsules. Furthermore, the swelling of pristine beads, pH-sensitive properties and in vitro release studies of curcumin loaded beads were investigated at 37 °C in simulated gastric fluid (SGF), simulated intestinal fluid (SIF) and simulated colonic fluid (SCF). The effect of the polymer blend ratio, the encapsulation efficiency (EE %) of curcumin, the loading capacity (LC µg/mg), the sphericity factor (SF), the antioxidant activity of the elaborated beads and their antimicrobial properties against bacteria and fungi were just as much evaluated. The obtained results indicate that the swelling and the behavior of the developed beads were influenced by the pH of the test medium and the PVA content. The introduction of PVA into the SA matrix greatly enhanced the physicochemical properties, the encapsulation efficiency and the loading capacity of the elaborated microparticles. Results also suggested that the antioxidant activity of the loaded beads (CA/PVA/Cur) showed a higher DPPH radical scavenging activity while the bacterial and fungal strains proved sensitive to the different formulations used in the assay. Moreover, the important drug encapsulation efficiency and the sustainable drug release of these materials make them promising for the development of new drug carrier systems for colon targeting.

Generation of Endotoxin-Specific Monoclonal Antibodies by Phage and Yeast Display for Capturing Endotoxin.

Endotoxin, a synonym for lipopolysaccharide (LPS), is anchored in the outer membranes of Gram-negative bacteria. Even minute amounts of LPS entering the circulatory system can have a lethal immunoactivating effect. Since LPS is omnipresent in the environment, it poses a great risk of contaminating any surface or solution, including research products and pharmaceuticals. Therefore, monitoring LPS contamination and taking preventive or decontamination measures to ensure human safety is of the utmost importance. Nevertheless, molecules used for endotoxin detection or inhibition often suffer from interferences, low specificity, and low affinity. For this reason, the selection of new binders that are biocompatible, easy to produce, and that can be used for biopharmaceutical applications, such as endotoxin removal, is of high interest. Powerful techniques for selecting LPS-binding molecules in vitro are display technologies. In this study, we established and compared the selection and production of LPS-specific, monoclonal, human single-chain variable fragments (scFvs) through two display methods: yeast and phage display. After selection, scFvs were fused to a human constant fragment crystallizable (Fc). To evaluate the applicability of the constructs, they were conjugated to polystyrene microbeads. Here, we focused on comparing the functionalized beads and their LPS removal capacity to a polyclonal anti-lipid A bead. Summarized, five different scFvs were selected through phage and yeast display, with binding properties comparable to a commercial polyclonal antibody. Two of the conjugated scFv-Fcs outperformed the polyclonal antibody in terms of the removal of LPS in aqueous solution, resulting in 265 times less residual LPS in solution, demonstrating the potential of display methods to generate LPS-specific binding molecules.

Surgical Reconstruction of Canine Nonunion Fractures Using Bone Morphogenetic Protein-2-loaded Alginate Microbeads and Bone Allografts.

BACKGROUND/AIM: Effective treatment of nonunion fractures is challenging as it requires a biological and mechanical environment to promote sufficient osteogenesis. Herein, we present a case series in which we evaluated the clinical efficacy of bone morphogenetic protein-2 (BMP-2)-loaded alginate microbeads and allografts in two dogs with nonunion fractures. CASE REPORT: A 3-year-old, 2.3-kg, spayed female Pomeranian (Case 1) presented with intermittent lameness of the left forelimb after radial and ulnar fracture repair 8 weeks prior. A 4-year-old, 4.8-kg, spayed female Pomeranian (Case 2) was referred for non-weight-bearing lameness of the left hindlimb due to implant failure following left tibial fracture repair. Both dogs had atrophic bone ends and no bridging calluses at the fracture site on radiographs, and were diagnosed with nonviable nonunion fractures of the radius/ulna and tibia, respectively. The surgical approach involved implant removal, debridement, and fracture gap reconstruction. BMP-2 was loaded into alginate microbeads for a prolonged release with bone allograft chips in both cases. In Case 1, bead grafts were applied directly at the fracture site, while in Case 2, they were implanted inside a frozen cortical bone allograft as a scaffold to fill the large gap. Postoperative radiography revealed excessive callus formation, early radiographic bone union, and cortical bone remodeling, in line with improved lameness scores. At the final follow-up, gait was improved and the desired bone length and shape were achieved in both cases. CONCLUSION: Simultaneous use of osteoinductive BMP-2 alginate microbeads and osteoconductive bone allografts yielded functionally and structurally favorable outcomes in canine nonunion fractures, without major complications.

Fabrication, characterization and release behavior of α-tocopherol acetate-loaded pH-responsive polyhydroxybutyrate/cellulose acetate phthalate microbeads.

Microbeads are used in personal care and cosmetic products (PCCPs) but are produced from nondegradable materials. Biodegradable polyhydroxybutyrate (PHB) has been recognized as a promising alternative material for use in PCCPs; however, utilizing PHB to encapsulate PCCPs is challenging because PCCPs need to be protected from the environment but their release needs to be permitted under specific physiological conditions. The aim of this work was to develop and evaluate pH-responsive cellulose acetate phthalate (CAP) to formulate lipophilic α-tocopherol acetate (α-TA)-loaded pH-responsive PHB/CAP microbeads. The influences of the PHB/CAP ratio and initial α-TA loading on the microbead size, surface morphology, encapsulation efficiency (%EE), loading capacity (%LC), and α-TA release profile were studied. The microbeads exhibited a spherical shape with a size of 328.7 ± 2.9 µm. The EE and LC were 86.7 ± 2.6 % and 13.5 ± 0.4 %, respectively. The release profile exhibited pH-responsive characteristics. These α-TA-loaded pH-responsive microbeads were stable with >50 % of the α-TA remaining after 90 days at 4, 25 and 45 °C in the dark. The results from the cytotoxicity assay with PSVK1 cells demonstrated that the microbeads were nontoxic. Hence, our developed formulation has the potential to be used to encapsulate oil-based drugs to formulate lipophilic substance-loaded pH-responsive microbeads.