In hybrid gels with immobilized liquid crystal
(LC) droplets, fast and unique optical texture variations are
generated when distinct volatile organic compounds (VOCs)
interact with the LC and disturb its molecular order. The
optical texture variations can be observed under a polarized
optical microscope or transduced into a signal representing the
variations of light transmitted through the LC. We show how
hybrid gels can accurately identify 11 distinct VOCs by using
deep learning to analyze optical texture variations of individual
droplets (0.93 average F1-score) and by using machine learning
to analyze 1D optical signals from multiple droplets in hybrid
gels (0.88 average F1-score)

We report the development of gas-sensing multicomponent hybrid materials to be used under humidified and dried environments without the need of sample preconditioning or heavy signal processing. The easy tunability and the unique characteristics presented by the multicomponent hybrid materials suggests their use in nearterm applications in electronic nose systems able to operate in dry or humidified environments.

Electronic noses (e-noses) mimic human olfaction, by identifying Volatile Organic Compounds (VOCs). This
work presents a novel approach that successfully classifies 11 known VOCs using the signals generated by
sensing gels in an in-house developed e-nose. The proposed signals’ analysis methodology is based on the
generated signals’ morphology for each VOC since different sensing gels produce signals with different shapes
when exposed to the same VOC. For this study, two different gel formulations were considered, and an average
f1-score of 84% and 71% was obtained, respectively. Moreover, a standard method in time series classification
was used to compare the performances. Even though this comparison reveals that the morphological approach
is not as good as the 1-nearest neighbour with euclidean distance, it shows the possibility of using descriptive
sentences with text mining techniques to perform VOC classification.

Relative humidity is simultaneously a sensing target and a contaminant in gas and volatile organic compound (VOC) sensing systems, where strategies to control humidity interference are required. An unmet challenge is the creation of gas-sensitive materials where the response to humidity is controlled by the material itself. Here, humidity effects are controlled through the design of gelatin formulations in ionic liquids without and with liquid crystals as electrical and optical sensors, respectively. In this design, the anions [DCA]− and [Cl]− of room temperature ionic liquids from the 1-butyl-3-methylimidazolium family tailor the response to humidity and, subsequently, sensing of VOCs in dry and humid conditions. Due to the combined effect of the materials formulations and sensing mechanisms, changing the anion from [DCA]− to the much more hygroscopic [Cl]−, leads to stronger electrical responses and much weaker optical responses to humidity. Thus, either humidity sensors or humidity-tolerant VOC sensors that do not require sample preconditioning or signal processing to correct humidity impact are obtained. With the wide spread of 3D- and 4D-printing and intelligent devices, the monitoring and tuning of humidity in sustainable biobased materials offers excellent opportunities in e-nose sensing arrays and wearable devices compatible with operation at room conditions.

Electronic noses (e-noses) mimic the complex biological olfactory system, usually including an array of gas sensors to act as the olfactory receptors and a trained computer with signal-processing and pattern recognition tools as the brain. In this work, a new stimuli-responsive material is shown, consisting of self-assembled droplets of liquid crystal and ionic liquid stabilised within a fish gelatin matrix. These materials change their opto/electrical properties upon contact with volatile organic compounds (VOCs). By using an in-house developed e-nose, these new gas-sensing films yield characteristic optical signals for VOCs from different chemical classes. A support vector machine classifier was implemented based on 12 features of the signals. The results show that the films are excellent identifying hydrocarbon VOCs (toluene, heptane and hexane) (95% accuracy) but lower performance was found to other VOCs, resulting in an overall 60.4% accuracy. Even though they are not reusable, these sustainable gas-sensing films are stable throughout time and reproducible, opening several opportunities for future optoelectronic devices and artificial olfaction systems.

The fast and non-invasive detection of odors and volatile organic compounds (VOCs) by gas sensors and electronic
noses is a growing field of interest, mostly due to a large scope of potential applications. Additional drivers for the
expansion of the field include the development of alternative and sustainable sensing materials. The discovery
that isolated cross-linked polymeric structures of suberin spontaneously self-assemble as a film inspired us to
develop new sensing composite materials consisting of suberin and a liquid crystal (LC). Due to their stimuliresponsive and optically active nature, liquid crystals are interesting probes in gas sensing. Herein, we report
the isolation and the chemical characterization of two suberin types (from cork and from potato peels) resorting to
analyses of gas chromatography–mass spectrometry (GC-MS), solution nuclear magnetic resonance (NMR), and Xray photoelectron spectroscopy (XPS). The collected data highlighted their compositional and structural differences. Cork suberin showed a higher proportion of longer aliphatic constituents and is more esterified than potato
suberin. Accordingly, when casted it formed films with larger surface irregularities and a higher C/O ratio. When
either type of suberin was combined with the liquid crystal 5CB, the ensuing hybrid materials showed distinctive
morphological and sensing properties towards a set of 12 VOCs (comprising heptane, hexane, chloroform,
toluene, dichlormethane, diethylether, ethyl acetate, acetonitrile, acetone, ethanol, methanol, and acetic acid).
The optical responses generated by the materials are reversible and reproducible, showing stability for 3 weeks.
The individual VOC-sensing responses of the two hybrid materials are discussed taking as basis the chemistry of
each suberin type. A support vector machines (SVM) algorithm based on the features of the optical responses was
implemented to assess the VOC identification ability of the materials, revealing that the two distinct suberin-based
sensors complement each other, since they selectively identify distinct VOCs or VOC groups. It is expected that
such new environmentally-friendly gas sensing materials derived from natural diversity can be combined in arrays
to enlarge selectivity and sensing capacity.

The materials described in this work result from the selfassembly of liquid crystals and ionic liquids into droplets,
stabilized within a biopolymeric matrix. These systems are
extremely versatile gels, in terms of composition, and offer
potential for fine tuning of both structure and function, as
each individual component can be varied. Here, the
characterization and application of these gels as sensing thin
films in gas sensor devices is presented. The unique
supramolecular structure of the gels is explored for molecular
recognition of volatile organic compounds (VOCs) by
employing gels with distinct formulations to yield
combinatorial optical and electrical responses used in the
distinction and identification of VOCs.

Affinity‐triggered assemblies rely on affinity interactions as the driving force to assemble physically‐crosslinked networks. WW domains are small hydrophobic proteins binding to proline‐rich peptides that are typically produced in the insoluble form. Previous works attempted the biological production of the full WW domain in tandem to generate multivalent components for affinity‐triggered hydrogels. In this work, an alternative approach was followed by engineering a 13‐mer minimal version of the WW domain that retains the ability to bind to target proline‐rich peptides. Both ligand and target peptides were produced chemically and conjugated to multivalent polyethylene glycol, yielding two components. Upon mixing, they together form soft biocompatible affinity‐triggered assemblies, stable in stem cell culture media, and displaying mechanical properties in the same order of magnitude as for those hydrogels formed with the full WW protein in tandem.

Artificial olfaction is a fast-growing field aiming to mimic natural olfactory systems. Olfactory systems rely on a first step of molecular recognition in which volatile organic compounds (VOCs) bind to an array of specialized olfactory proteins. This results in electrical signals transduced to the brain where pattern recognition is performed. An efficient approach in artificial olfaction combines gas-sensitive materials with dedicated signal processing and classification tools. In this work, films of gelatin hybrid gels with a single composition that change their optical properties upon binding to VOCs were studied as gas-sensing materials in a custom-built electronic nose. The effect of films thickness was studied by acquiring signals from gelatin hybrid gel films with thicknesses between 15 and 90 μm when exposed to 11 distinct VOCs. Several features were extracted from the signals obtained and then used to implement a dedicated automatic classifier based on support vector machines for data processing. As an optical signature could be associated to each VOC, the developed algorithms classified 11 distinct VOCs with high accuracy and precision (higher than 98%), in particular when using optical signals from a single film composition with 30 μm thickness. This shows an unprecedented example of soft matter in artificial olfaction, in which a single gelatin hybrid gel, and not an array of sensing materials, can provide enough information to accurately classify VOCs with small structural and functional differences.

Electronic noses (E-noses), are usually composed by an array of sensors with different selectivities towards classes of VOCs (Volatile Organic Compounds). These devices have been applied to a variety of fields, including environmental protection, public safety, food and beverage industries, cosmetics, and clinical diagnostics. This work demonstrates that it is possible to classify eleven VOCs from different chemical classes using a single gas sensing biomaterial that changes its optical properties in the presence of VOCs. To accomplish this, an in-house built E-nose, tailor-made for the novel class of gas sensing biomaterials, was improved and combined with powerful machine learning techniques. The device comprises a delivery system, a detection system and a data acquisition and control system. It was designed to be stable, miniaturized and easy-to-handle. The data collected was pre-processed and features and curve fitting parameters were extracted from the original response. A recursive feature selection method was applied to select the best features, and then a Support Vector Machine classifier was implemented to distinguish the eleven distinct VOCs. The results show that the followed methodology allowed the classification of all the VOCs tested with 94.6% (± 0.9%) accuracy.

The cooperative assembly of biopolymers and small molecules can yield functional materials with precisely tunable properties. Here, the fabrication, characterization, and use of multicomponent hybrid gels as selective gas sensors are reported. The gels are composed of liquid crystal droplets self-assembled in the presence of ionic liquids, which further coassemble with biopolymers to form stable matrices. Each individual component can be varied and acts cooperatively to tune gels' structure and function. The unique molecular environment in hybrid gels is explored for supramolecular recognition of volatile compounds. Gels with distinct compositions are used as optical and electrical gas sensors, yielding a combinatorial response conceptually mimicking olfactory biological systems, and tested to distinguish volatile organic compounds and to quantify ethanol in automotive fuel. The gel response is rapid, reversible, and reproducible. These robust, versatile, modular, pliant electro-optical soft materials possess new possibilities in sensing triggered by chemical and physical stimuli.

Adenoviruses are important platforms for vaccine development and vectors for gene therapy, increasing the demand for high titers of purified viral preparations. Monoliths are macroporous supports regarded as ideal for the purification of macromolecular complexes, including viral particles. Although common monoliths are based on synthetic polymers as methacrylates, we explored the potential of biopolymers processed by clean technologies to produce monoliths for adenovirus purification. Such an approach enables the development of disposable and biodegradable matrices for bioprocessing. A total of 20 monoliths were produced from different biopolymers (chitosan, agarose, and dextran), employing two distinct temperatures during the freezing process (−20 °C and −80 °C). The morphological and physical properties of the structures were thoroughly characterized. The monoliths presenting higher robustness and permeability rates were further analyzed for the nonspecific binding of Adenovirus serotype 5 (Ad5) preparations. The matrices presenting lower nonspecific Ad5 binding were further functionalized with quaternary amine anion-exchange ligand glycidyltrimethylammonium chloride hydrochloride by two distinct methods, and their performance toward Ad5 purification was assessed. The monolith composed of chitosan and poly(vinyl) alcohol (50:50) prepared at −80 °C allowed 100% recovery of Ad5 particles bound to the support. This is the first report of the successful purification of adenovirus using monoliths obtained from biopolymers processed by clean technologies.

BACKGROUND The emergence of monoclonal antibodies (mAbs) as new biopharmaceutical products requires the development of new purification methods that are not only effective but are able to reduce production costs. To address the problematic recovery of mAbs, gum arabic (GA) coated magnetic particles (MPs) were used for the purification of human antibodies from animal cells supernatants. RESULTS MPs were synthesized via co-precipitation and exhibited a spherical-like physical aspect, with an average hydrodynamic diameter of 473 nm and a zeta potential of –26 mV. The adsorption and elution of IgG on these adsorbents was thoroughly studied. Adsorption of human IgG was enhanced at pH 6, for which a qmax of 244 mg IgG g−1 MPs and Kd of 25 mg L−1 were obtained. Increasing salt concentrations at a basic pH (1 mol L−1 NaCl at pH 11) were found to improve the elution of bound IgG. The MPs were challenged with an artificial protein mixture containing human IgG, albumin, insulin and apo-transferrin. An overall yield of 84% was achieved, retrieving 92% of bound IgG. CONCLUSIONS MPs were successfully used for the capture of monoclonal antibodies from two distinct mammalian cell cultures, a Chinese hamster ovary (CHO) and a hybridoma cell culture supernatants. The elution yields were high, ranging between 84% and 94%, with overall yields ranging from 72% to 88%. Final purities of 85% were reached for hybridoma cell supernatants. © 2014 Society of Chemical Industry

Aminophenyl boronic acids can form reversible covalent ester interactions with cis-diol-containing molecules, serving as a selective tool for binding glycoproteins as antibody molecules that possess oligosaccharides in both the Fv and Fc regions. In this study, amino phenyl boronic acid (APBA) magnetic particles (MPs) were applied for the magnetic separation of antibody molecules. Iron oxide MPs were firstly coated with dextran to avoid non-specific binding and then with 3-glycidyloxypropyl trimethoxysilane to allow further covalent coupling of APBA (APBA_MP). When contacted with pure protein solutions of human IgG (hIgG) and bovine serum albumin (BSA), APBA_MP bound 170 ± 10 mg hIgG g−1 MP and eluted 160 ± 5 mg hIgG g−1 MP, while binding only 15 ± 5 mg BSA g−1 MP. The affinity constant for the interaction between hIgG and APBA_MP was estimated as 4.9 × 105 M−1 (Ka) with a theoretical maximum capacity of 492 mg hIgG adsorbed g−1 MP (Qmax), whereas control particles bound a negligible amount of hIgG and presented an estimated theoretical maximum capacity of 3.1 mg hIgG adsorbed g−1 MP (Qmax). APBA_MPs were also tested for antibody purification directly from CHO cell supernatants. The particles were able to bind 98% of IgG loaded and to recover 95% of pure IgG (purity greater than 98%) at extremely mild conditions.

Industrial and urban activities yield large amounts of contaminated groundwater, which present a major health issue worldwide. Infectious diseases are the most common health risk associated with drinking-water and wastewater remediation is a major concern of our modern society. The field of wastewater treatment is being revolutionized by new nano-scale water disinfection devices which outperform most currently available technologies. In particular, iron oxide magnetic nanoparticles (MNPs) have been widely used in environmental applications due to their unique physical–chemical properties. In this work, poly(ethylene) glycol (PEG)-coated MNPs have been functionalized with (RW)3, an antimicrobial peptide, to yield a novel magnetic-responsive support with antimicrobial activity against Escherichia coli K-12 DSM498 and Bacillus subtilis 168. The magnetic-responsive antimicrobial device showed to be able to successfully disinfect the surrounding solution. Using a rapid high-throughput screening platform, the minimal inhibitory concentration (MIC) was determined to be 500 μM for both strains with a visible bactericidal effect.

FucoPol, a fucose-containing extracellular polysaccharide (EPS) produced by bacterium Enterobacter A47 using glycerol as the carbon source, was employed as a coating material for magnetic particles (MPs), which were subsequently functionalized with an artificial ligand for the capture of antibodies. The performance of the modified MPs (MP–EPS-22/8) for antibody purification was investigated using direct magnetic separation alone or combined with an aqueous two-phase system (ATPS) composed of polyethylene glycol (PEG) and dextran. In direct magnetic capturing, and using pure protein solutions of human immunoglobulin G (hIgG) and bovine serum albumin (BSA), MP–EPS-22/8 bound 120 mg hIgG g−1 MPs, whereas with BSA only 10 ± 2 mg BSA g−1 MPs was achieved. The hybrid process combining both the ATPS and magnetic capturing leads to a good performance for partitioning of hIgG in the desired phase as well as recovery by the magnetic separator. The MPs were able to bind 145 mg of hIgG g−1 of particles which is quite high when compared with direct magnetic separation. The theoretical maximum capacity was calculated to be 410 ± 15 mg hIgG adsorbed g−1 MPs with a binding affinity constant of 4.3 × 104 M−1. In multiple extraction steps, the MPs bound 92% of loaded hIgG with a final purity level of 98.5%. The MPs could easily be regenerated, recycled and re-used for five cycles with only minor loss of capacity. FucoPol coating allowed both electrostatic and hydrophobic interactions with the antibody contributing to enhance the specificity for the targeted products.

Monoliths represent powerful platforms for isolation of large molecules with high added value. This work presents a hybrid approach for antibody (Ab) capture and release. Using mostly natural polymers and clean processes, it is possible to create macroporous monoliths with well-defined porous networks, tuneable mechanical properties, and easy functionalization with a biomimetic ligand specific for Ab. Magnetic nanoparticles (MNPs) are embedded on the monolith network to confer a controlled magnetic response that facilitates and accelerates Ab recovery in the elution step. The hybrid monolithic systems prepared with agarose or chitosan/poly(vinyl alcohol) (PVA) blends exhibit promising binding capacities of Abs directly from cell-culture extracts (120 ± 10 mg Ab g−1 support) and controlled Ab magnetically-assisted elution yielding 95 ± 2% recovery. Moreover, a selective capture of mAbs directly from cell culture extracts is achieved yielding a final mAb preparation with 96% of purity.

The potential to combine aqueous two-phase extraction (ATPE) with magnetic separation was here investigated with the aim of developing a selective non-chromatographic method for the purification of antibodies from cell culture supernatants. Aqueous two-phase systems (ATPS) composed of polyethylene glycol (PEG) and dextran were supplemented with several surface modified magnetic particles (MPs) at distinct salt concentrations. The partition of pure human IgG in the upper and lower phases as well as the amount adsorbed at the MPs surface was investigated, indicating that MPs coated with dextran and gum Arabic established the lowest amount of non-specific interactions. The binding capacity of gum arabic coated particles modified with aminophenyl boronic acid (GA-APBA-MP) was were found to be excellent in combination with the ATPS system, yielding high yields of antibody recovery (92%) and purity (98%) from cell culture supernatants. The presence of MPs in the ATPS was found to speed up phase separation (from 40 to 25 min), to consume a lower amount of MPs (half of the amount needed in magnetic fishing) and to increase the yield and purity of a mAb purified from a cell culture supernatant, when compared with ATPE or magnetic fishing processes alone.

Affinity chromatography is one of the most common techniques employed at the industrial-scale for antibody purification. In particular, the purification of human immunoglobulin G (hIgG) has gained relevance with the immobilization of its natural binding counterpart—Staphylococcus aureus Protein A (SpA) or with the recent development of biomimetic affinity ligands, namely triazine-based ligands. These ligands have been developed in order to overcome economic and leaching issues associated to SpA. The most recent triazine-based ligand—TPN-BM, came up as an analogue of 2-(3-amino-phenol)-6-(4-amino-1-naphthol)-4-chloro-sym-triazine ligand also known as ligand 22/8 with improved physico-chemical properties and a greener synthetic route. This work intends to evaluate the potential of TPN-BM as an alternative affinity ligand towards antibody recognition and binding, namely IgG, at an atomic level, since it has already been tested, after immobilization onto chitosan-based monoliths and demonstrated interesting affinity behaviour for this purpose. Herein, combining automated molecular docking and molecular dynamics simulations it was predicted that TPN-BM has high propensity to bind IgG through the same binding site found in the crystallographic structure of SpA_IgG complex, as well as theoretically predicted for ligand 22/8_IgG complex. Furthermore, it was found that TPN-BM established preferential interactions with aromatic residues at the Fab domain (Trp 50, Tyr 53, Tyr 98 and Trp 100), while in the Fc domain the main interactions are based on hydrogen bonds with pH sensitive residues at operational regime for binding and elution like histidines (His 460, His 464, His 466). Moreover, the pH dependence of TPN-BM_IgG complex formation was more evident for the Fc domain, where at pH 3 the protonation state and consequently the charge alteration of histidine residues located at the IgG binding site induced ligand detachment which explains the optimal elution condition at this pH observed experimentally.

Polymer monoliths are an efficient platform for antibody purification. The use of monoclonal antibodies (mAbs) and engineered antibody structures as therapeutics has increased exponentially over the past few decades. Several approaches use polymer monoliths to purify large quantities of antibody with defined clinical and performance requirements. Functional monolithic supports have attracted a great deal of attention as they offer practical advantages for antibody purification, such as more rapid analysis, smaller sample volume requirements and the opportunity for a greater target molecule enrichment. This review focuses on the development of synthetic and natural polymer-based monoliths for antibody purification. The materials and methods employed in monolith production are discussed, highlighting the properties of each system. We also review the structural characterization techniques available using monolithic systems and their performance under different chromatographic approaches to antibody capture and release. Finally, a summary of monolithic platforms developed for antibody separation is presented, as well as expected trends in research to solve current and future challenges in this field. This review comprises a comprehensive analysis of proposed solutions highlighting the remarkable potential of monolithic platforms.

Magnetic separations are probably one of the most versatile separation processes in biotechnology as they are able to purify cells, viruses, proteins and nucleic acids directly from crude samples. The fast and gentle process in combination with its easy scale-up and automation provide unique advantages over other separation techniques. In the midst of this process are the magnetic adsorbents tailored for the envisioned target and whose complex synthesis spans over multiple fields of science. In this context, this article reviews both the synthesis and tailoring of magnetic adsorbents for bioseparations as well as their ultimate application.

Monoclonal antibodies (mAbs) are important therapeutic proteins. One of the challenges facing large-scale production of monoclonal antibodies is the capacity bottleneck in downstream processing, which can be circumvented by using magnetic stimuli-responsive polymer nanoparticles. In this work, stimuli-responsive magnetic particles composed of a magnetic poly(methyl methacrylate) core with a poly(N-isopropylacrylamide-co-acrylic acid) (P(NIPAM-co-AA)) shell cross-linked with N, N'-methylenebisacrylamide were prepared by miniemulsion polymerization. The particles were shown to have an average hydrodynamic diameter of 317 nm at 18°C, which decreased to 277 nm at 41°C due to the collapse of the thermo-responsive shell. The particles were superparamagnetic in behavior and exhibited a saturation magnetization of 12.6 emu/g. Subsequently, we evaluated the potential of these negatively charged stimuli-responsive magnetic particles in the purification of a monoclonal antibody from a diafiltered CHO cell culture supernatant by cation exchange. The adsorption of antibodies onto P(NIPAM-co-AA)-coated nanoparticles was highly selective and allowed for the recovery of approximately 94% of the mAb. Different elution strategies were employed providing highly pure mAb fractions with host cell protein (HCP) removal greater than 98%. By exploring the stimuli-responsive properties of the particles, shorter magnetic separation times were possible without significant differences in product yield and purity.

Chitosan-based monoliths activated by plasma technology induced the coupling of a robust biomimetic ligand, previously reported as an artificial Protein A, with high yields while minimizing the environmental impact of the procedure. Due to the high porosity, good mechanical and tunable physicochemical properties of the affinity chitosan-based monoliths, it is possible to achieve high binding capacities (150 ± 10 mg antibody per gram support), and to recover 90 ± 5% of the bound protein with 98% purity directly from cell-culture extracts. Therefore, the chitosan-based monoliths prepared by clean processes exhibit a remarkable performance for the one-step capture and recovery of pure antibodies or other biological molecules with biopharmaceutical relevance.

In this work we have evaluated the potential of boronic acid functionalized magnetic particles for the one-step capture of a human monoclonal antibody (mAb) from a Chinese hamster ovary (CHO) cell culture supernatant. For comparison, Protein A coated magnetic particles were also used. The most important factor influencing the overall process yield and product purity in boronic acid particles was found to be the binding pH. Basic pH values promoted higher purities while resulting in decreased yields due to the competing effects of molecules such as glucose and lactate present in the cell culture supernatant. After optimization, the particles were successfully used in a multi-cycle purification process of the mAb from the CHO feedstock. Boronic acid particles were able to achieve an average overall yield of 86% with 88% removal of CHO host cell proteins (HCP) when the binding was performed at pH 7.4, while at pH 8.5 these values were 58% and 97%, respectively. In both cases, genomic DNA removal was in excess of 97%. Comparatively, Protein A particles recorded an average overall yield of 80% and an HCP removal greater than 99%. The adsorption of the mAb to the boronic acid particles was shown to be mediated by strong affinity interactions. Overall, boronic acid based purification processes can offer a cost-effective alternative to Protein A as the direct capturing step from the mammalian cell culture.

Fluorescence microscopy and microspectrofluorometry are important tools in the characterization and identification of proteins, offering a great range of applications in conservation science. Because of their high selectivity and sensitivity, the combination of these techniques can be exploited for improved recognition and quantification of proteinaceous binders in paintings and polychromed works of art. The present article explores an analytical protocol integrating fluorescence microscopy and fluorometry for both identification and mapping of proteinaceous binders (in particular egg and glues) in paint samples. The study has been carried out on historically accurate reconstructions simulating the structure and composition of tempera and oil paints containing these binders. To assess the spatial distribution of specific proteins within the paint layers, cross-sections from the reconstructions were analyzed by fluorescence imaging after staining with an exogenous fluorophore. Reference fluorescence spectra for each layer were acquired by a multichannel spectral analyzer and compared after Gaussian deconvolution. The results obtained demonstrated the effectiveness of the integrated protocol, highlighting the potential for the use of fluorescent staining coupled with microspectrofluorometry as a routine diagnostic tool in conservation science. The current work creates a set of fully characterized reference samples for further comparison with those from actual works of art.

In this work, we systematically evaluated the potential of using boronic acid functionalized magnetic particles in the capturing of human immunoglobulin G under typical mammalian cell culture conditions. For comparison, Protein A coated magnetic particles were also used. The binding pH was found to significantly influence the adsorption isotherms of boronic acid particles with the higher capacities (0.216 g IgG/g support) being observed at pH 7.4. Comparatively, this value was 0.109 g IgG/g support, for Protein A particles under the same conditions. Both particles revealed very fast adsorption kinetics with more than 70% of the maximum binding capacity being achieved in a few seconds. The effect of glucose and lactate, which are known to interact with boronic acid, was evaluated. For glucose, the binding capacity was significantly influenced by the pH and decreased as pH increased. At pH 9.5, a 70% lower binding capacity was observed for glucose concentrations as low as 0.5 g/l. The effect of lactate was less pronounced and almost pH independent reaching at most 20% decrease in binding capacity. Nevertheless, the effect of both molecules was always lower at pH 7.4. The optimization of the elution conditions enabled complete recovery of bound IgG from boronic acid particles using 50mM Tris-HCl, 200 mM sorbitol, 200 mM NaCl at pH 8.5.

This paper reports the design, preparation and characterization of cellulose affinity membranes for antibody purification using a new methodology. Cellulose membranes were prepared from polymer-ionic liquid solutions, namely 1-butyl-3-methylimidazolium chloride {([BMIM][Cl])}, by the water induced phase inversion process. After functionalization with a synthetic ligand 2-(3-aminophenol)-6-(4-amino-1-naphthol)-4-chloro-s-triazine (ligand 22/8), these were evaluated as affinity supports for human immunoglobulin G {(IgG).} Membranes were characterized in terms of morphology {(SEM)}, porosity (mercury porosimetry), hydrophilicity (contact angle measurement), transport properties (permeability) and mechanical performance {(DMA).} Membranes prepared with varying cellulose contents (5 and 10&\#xa0;wt.% cellulose in ionic liquid solutions) lead to films with different properties. The 10&\#xa0;wt.% cellulose membrane presented enhanced morphological and mechanical properties, however, the morphology of this membrane was significantly altered after ligand coupling. Adsorption isotherms for human {IgG} onto 10&\#xa0;wt.% matrix activated with ligand 22/8 were obtained. Preliminary results showed that the bovine serum albumin {(BSA)}, a model impurity, did not adsorb onto the membrane while up to 6&\#xa0;mg {IgG/g} was bound and 2&\#xa0;mg {IgG/g} recovered.

Magnetic particles {(MNPs)} offer attractive possibilities in biotechnology. {MNPs} can get close to a target biological entity, as their controllable sizes range from a few nanometres up to tens of nanometres, and their surface can be modified to add affinity and specificity towards desired molecules. Additionally, they can be manipulated by an external magnetic field gradient. In this work, the study of ferric oxide {(Fe3O4)} {MNPs} with different coating agents was conducted, particularly in terms of strategies for antibody attachment at the surfaces (covalent and physical adsorption) and the effects of blocking buffer composition and incubation times on the specific and non-specific interactions observed. The considered biological model system consisted of a coating antibody (goat {IgG)}, bovine serum albumin {(BSA)} as blocking agent, and a complementary antibody labelled with {FITC} (anti-goat {IgG).} The detection of antibody binding was followed by fluorescence microscopy and the intensity of the signals quantified. The ratio between the mean grey values of negative and positive controls, as well as the maximum intensity attainable in positive controls, were considered in the evaluation of the assays efficiency. The covalent immobilization of the coating antibody was more successful as opposed to protein adsorption. For covalent immobilization, silica-coated {MNPs}, a 5% (w/v) concentration of {BSA} in the blocking buffer and incubation times of 1 h produced the best results in terms of assay sensitivity. However, when conducting the assay for incubation periods of 10 min, the fluorescence signal was reduced by 44% but the assay specificity was maintained.