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Soil contamination by multiple metals is a significant concern due to the interlinked mobilization processes. The challenges in comprehending this issue arise from the poorly characterized interaction among different metals and the complexities introduced by spatial and temporal heterogeneity in soil systems. We delved into these complexities by incubating size-fractionated paddy soils under both anaerobic and aerobic conditions, utilizing a combination of techniques for aqueous and colloidal analysis. The contaminated paddy soil predominantly consisted of particles measuring <53, 250-53, and 2000-250 μm, with the <53 μm fractions exhibiting the highest concentrations of multiple metals. Interestingly, despite their higher overall content, the <53 μm fractions released less dissolved metal. Furthermore, glucose enhanced the release of arsenic while simultaneously promoting the sequestration of other metals, such as Pb, Zn, and Cu. Utilizing asymmetric flow field-flow fractionation, we unveiled the presence of both fine (0.3-130 kDa) and large (130-450 nm) colloidal pools, each carrying various metals with different affinities for iron minerals and organic matter. Our results highlighted the pivotal role of the <53 μm fraction as a significant reservoir for multiple metal contaminants in paddy soils, in which the colloidal metals were mainly associated with organic matter. These findings illuminated the size-resolved dynamics of soil metal cycling and provided insights for developing remediation strategies for metal-contaminated soil ecosystems.

Nanomedicines comprise multiple components, and particle density is considered an important property that regulates the biodistribution of administered nanomedicines. The density of nanoparticles is characterized by centrifugal methods, such as analytical ultracentrifugation. Particle size and distribution are key physicochemical and quality attributes of nanomedicines. In this study, we developed a novel profiling method applicable to liposomes and lipid nanoparticles (LNPs), based on particle size and density, using centrifugal field-flow fractionation (CF3). We evaluated the elution profiles of PEGylated liposomes of different sizes with various doxorubicin (DOX)-loading amounts using CF3. This method was applied to evaluate the drug release of DOX-loaded liposomes, intra- and inter-batch variability, reconstitution reproducibility of AmBisome®, and elution characteristics of LNPs in COVID-19 vaccines (Comirnaty® and Spikevax TM ). The data obtained in the present study underscore the significance of the proposed methodology and highlight the importance of profiling and characterizing liposomes and LNPs using CF3 fractograms and a multi-angle light-scattering detector., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: [Eiichi Yamamoto reports financial support was provided by Japan Agency for Medical Research and Development. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper]., (Copyright © 2024 Elsevier B.V. All rights reserved.)

This study investigated critical metal (CM) geochemistry including rare earth elements (REEs), Co, Ni, and Mn in groundwaters below and surrounding two dredged material placement facilities (DMPFs). Metal concentrations are elevated at both sites, spanning several orders of magnitude. The highest CM concentrations measured exceed many environments considered as aqueous resources (Co and Ni > 1 mg L -1 , REEs > 3 mg L -1 ). Correlations between sulfur and iron, major cations, and CMs indicate that oxidation of sulfides present in the DM releases metals both directly from sulfide minerals and indirectly through acid dissolution of and/or desorption from additional minerals. REE fractionation patterns indicate that their mobility in the groundwaters may be influenced by interactions with silicate, carbonate, and phosphate minerals. Significant positive Gd and Eu anomalies were observed, which may be attributed to increased mobility of Eu 2+ and anthropogenic Gd. Nanogeochemical analysis of filtered samples revealed several REE-bearing nanoparticulate (diameter < 100 nm) species, some of which co-occurred with aluminum, suggesting an (oxy)hydroxide or a clay mineral component. Further characterization of soluble and nano scale geochemical speciation is needed to fully assess the viability of CM recovery from DM-associated groundwater. CM recovery from DM-associated waters can provide a beneficial use, both offsetting costs associated with disposal, and supplementing domestic CM resources., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Extracellular vesicles are small membrane-bound structures that are released by cells and play important roles in intercellular communication garnering significant attention in scientific society recently due to their potential as diagnostic and therapeutic tools. However, separating EVs from large-volume samples remains a challenge due to their small size and low concentration. In this manuscript, we presented a novel method for separating polystyrene beads as control and extracellular vesicles from large sample volumes using bifurcated asymmetric field flow fractionation in PDMS-free microfluidic devices. Separation characteristics were evaluated using the control system of polystyrene bead mix, which offers up to 3.7X enrichment of EV-sized beads. Furthermore, in the EV-sample from bioreactor culture media, we observed a notable population distribution shift of extracellular vesicles. Herein presented novel PDMS-free microfluidic device fabrication protocol resulted in devices with reduced EV-loss compared to size-exclusion columns. This method represented an improvement over the current state of the art in terms of EV separation from large sample volumes through the use of novel field flow fractionation design.

While the exact health risks associated with nanoplastics are currently the focus of intense research, there is no doubt that humans are exposed to nanoplastics and that food could be a major source of exposure. Nanoplastics are released from plastic materials and articles used during food production, processing, storage, preparation, and serving. They are also likely to enter the food chain via contaminated water, air, and soil. However, very limited exposure data for risk assessment exists so far due to the lack of suitable analytical methods. Nanoplastic detection in food poses a great analytical challenge due to the complexity of plastics and food matrices as well as the small size and expectedly low concentration of the plastic particles. Multidetector field flow fractionation has emerged as a valuable analytical technique for nanoparticle separation over the last decades, and the first studies using the technique for analyzing nanoplastics in complex matrices are emerging. In combination with online detectors and offline analysis, multidetector field flow fractionation is a powerful platform for advanced characterization of nanoplastics in food by reducing sample complexity, which otherwise hampers the full potential of most analytical techniques. The focus of this article is to present the current state of the art of multidetector field flow fractionation for nanoplastic analysis and to discuss future trends and needs aiming at the analysis of nanoplastics in food., (© 2022. Springer-Verlag GmbH Germany, part of Springer Nature.)

Hyaluronic acid (HA)-based prodrugs bearing double-responsive (acid pH or oxidation) boronates of catechol-containing drugs were used to treat xenografted human prostate tumours (LNCaP) in SCID mice. The HA prodrugs accumulated significantly only in tumours (impressively, up to 40% of the injected dose after 24 h) and in liver, with negligible - actually anti-inflammatory - consequences in the latter. A quercetin-HA prodrug significantly slowed down tumour growth, in a dose-dependent fashion and with a much higher efficacy (up to 4 times) than equivalent doses of free quercetin. In short, boronated HA appears to be a very promising platform for targeted chemotherapy., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Fluorescent probes are used in drug nanocarrier pre-clinical studies or as active compounds in theranostics and photodynamic therapy. In the biological medium, nanoparticles interact with proteins, which can result in the off-target release of their cargo. The present study used asymmetric flow field-flow fractionation with online multi-angle laser light scattering and fluorescence detection (AF4-MALLS-FLD) to study the release, transfer, and partition of fluorescent dyes from polymeric nanoparticles (NP). NP formulations containing the dyes Rose Bengal, Rhodamine B, DiI, 3-(α-azidoacetyl)coumarin and its polymer conjugate, Nile Red, and IR780 and its polymer conjugate were prepared. NP suspensions were incubated in a medium with serum proteins and then analyzed by AF4. AF4 allowed efficient separation of proteins (< 10 nm) from fluorescently labeled NP (range of 54 - 180 nm in diameters). The AF4 analyses showed that some dyes, such as Rose Bengal, IR780, and Coumarin were transferred to a high extent (68-77%) from NP to proteins. By contrast, for DiI and dye-polymer conjugates, transfer occured to a lower extent. The studies of dye release kinetics showed that the transfer of IR780 from NP to proteins occurs at a high extent (~50%) and rate, while Nile Red was slowly released from the NP over time with reduced association with proteins (~20%). This experiment assesses the stability of fluorescence labeling of nanocarriers and probes the transfer of fluorescent dyes from NP to proteins, which is otherwise not accessible with commonly used techniques of separation, such as dialysis and ultrafiltration/centrifugation employed in drug encapsulation and release studies of nanocarriers. Determining the interaction and transfer of dyes to proteins is of utmost importance in the pre-clinical evaluation of drug nanocarriers for improved correlation between in vitro and in vivo studies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Broad diversity and heterogeneity are inherently showcased by both natural and synthetic macromolecular structures. The high application potential for such structures and their combinations calls for novel analytical approaches that allow for comprehensive characterization and a full understanding of their complex composition. This review gives an overview of recent advances in designing and fabricating bioconjugated and self-assembled polymer structures, and introduces adequate characterization protocols for sufficient elucidation of their specific molecular properties. Possible pitfalls in their analysis are demonstrated, and potential alternatives are discussed. The primary focus is on addressing the highlights, and future prospects of applying field-flow fractionation coupled and/or hyphenated to different detection methods as a powerful separation and analytical technique for bioconjugate and self-assembled nanostructures., (© 2021 The Authors. Analytical Science Advances published by Wiley‐VCH GmbH.)

An analytical challenge that arises in environmental and food analysis is to quantify heterogeneous nanoparticles especially in polydisperse and complex samples. The method stated herein based on the reinjection asymmetrical flow field-flow fractionation (AF4 × AF4) coupled with inductively coupled plasma-mass spectrometer (ICP-MS) and statistical deconvolution allowed for identifying the molecular weight (Mw) and selenium abundance of the low Mw protein fractions (ca. < 132 kDa) in an unknown and complex sample (e.g., selenium-rich soybean protein isolates (Se-SPI)). A non-linear decay crossflow program was also developed to get better resolution and shorter elution time for both low and high Mw components. The concept of the reinjection method was based on the excellent ability for reducing sample complexity regarding the size fractionation, and peak reproducibility under the identical conditions of AF4 system. The standard protein mixture was used as a proof-of-principle sample. The results showed the underlying peaks predicted by the reinjection method were agreed with the separation result using the standard mixture (the relative standard deviation of peak locations < 1%), which indicated the reinjection method could provide an accurate assessment of the underlying peak number and location, and was promising to minimize the overfitting problem for statistic deconvolution. Interestingly, significant differences of Se abundance in protein fractions were observed in the low Mw range for Se-SPI, ranging from 0.28 to 1.66 cps/V with the Mw ranging from 13.75 kDa to 104.17 kDa, which indicated significant differences in the ability of binding Se for these selenium-rich proteins in Se-SPI., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier B.V.)

Dielectrophoresis-field flow fractionation (DEP-FFF) has emerged as an efficient in-vitro, non-invasive, and label-free mechanism to manipulate a variety of nano- and micro-scaled particles in a continuous-flow manner. The technique is mainly used to fractionate particles/cells based on differences in their sizes and/or dielectric properties by employing dielectrophoretic force as an external force field applied perpendicular to the flow direction. The dielectrophoretic force is the result of a spatially non-uniform electric field in the microchannel that can be generated either by exploiting microchannel geometry or using special arrangements of microelectrode arrays. Several two-dimensional (e.g., coplanar interdigitated, castellated) and three-dimensional (e.g., top-bottom, side-wall) microelectrode designs have been successfully utilized to perform fractionation of heterogeneous samples. Although originally introduced as a separation technique, DEP-FFF has attracted increasing interest in performing other important operations such as switching, focusing, dipping, and surface functionalization of target particles. Nonetheless, the technique still suffers from limitations such as low throughput and joule heating. By comparatively analyzing recent developments that address these shortcomings, this work is a step forward towards realizing the full potential of DEP-FFF as an ideal candidate for point-of-care (POC) devices with diverse applications in the fields of biomedical, chemical, and environmental engineering., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2020. Published by Elsevier B.V.)

Asymmetric-flow field-flow fractionation (AF4) has been recognized as an invaluable tool for the characterisation of particle size, polydispersity, drug loading and stability of nanopharmaceuticals. However, the application of robust and high quality standard operating procedures (SOPs) is critical for accurate measurements, especially as these complex drug nanoformulations are most often inherently polydisperse. In this review we describe a unique international collaboration that lead to the development of a robust SOP for the measurement of physical-chemical properties of nanopharmaceuticals by multi-detector AF4 (MD-AF4) involving two state of the art infrastructures in the field of nanomedicine, the European Union Nanomedicine Characterization Laboratory (EUNCL) and the National Cancer Institute-Nanotechnology Characterisation Laboratory (NCI-NCL). We present examples of how MD-AF4 has been used for the analysis of key quality attributes, such as particle size, shape, drug loading and stability of complex nanomedicine formulations. The results highlight that MD-AF4 is a very versatile analytical technique to obtain critical information on a material particle size distribution, polydispersity and qualitative information on drug loading. The ability to conduct analysis in complex physiological matrices is an additional very important advantage of MD-AF4 over many other analytical techniques used in the field for stability studies. Overall, the joint NCI-NCL/EUNCL experience demonstrates the ability to implement a powerful and highly complex analytical technique such as MD-AF4 to the demanding quality standards set by the regulatory authorities for the pre-clinical safety characterization of nanomedicines., Competing Interests: Declaration of Competing Interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020. Published by Elsevier B.V.)

Aquatic ecosystems are facing multiple direct and indirect stressors from anthropogenic activities. Aquatic colloids, including natural and manufactured nanoparticles, have received global concern owing to their ability to transport metals, nutrients and pollutants, and their potential to pose ecological risks for wildlife and human health. Asymmetric flow-field flow fractionation (AF4) has become one of the most promising and powerful approaches to characterize colloidal particles over the last ten years. This review overviews the advances and applications of AF4 for the study of aquatic systems over the last decade. The analytical challenge of AF4 and known deviation from theoretical assumptions are also critically discussed, such as sample losses, membrane-analyte interactions, and overloading. Future needs for methodological advances are addressed., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier B.V.)

Biogeochemical changes of whole catchments may, at least in part, be deduced from changes in stream water composition. We hypothesized that there are seasonal variations of natural nanoparticles (NNP; 1-100 nm) and fine colloids (<300 nm) in stream water, which differ in origin depending on catchment inflow parameters. To test this hypothesis, we assessed the annual dynamics of the elemental composition of NNP and fine colloids in multiple water compartments, namely in stream water, above and below canopy precipitation, groundwater and lateral subsurface flow from the Conventwald catchment, Germany. In doing so, we monitored meteorological and hydrological parameters, total element loads, and analyzed element concentrations of org C, Al, Si, P, Ca, Mn and Fe by Asymmetric Flow Field Flow Fractionation (AF 4 ). The results showed that colloid element concentrations were < 5 µmol/L. Up to an average of 55% (Fe) of total element concentrations were not truly dissolved but bound to NNP and fine colloids. The colloid patterns showed seasonal variability with highest loads in winter. The presence of groundwater-derived colloidal Ca in stream water showed that groundwater mainly fed the streams throughout the whole year. Overall, the results showed that different water compartments vary in the NNP and fine colloidal composition making them a suitable tool to identify the streams NNP and fine colloid sources. Given the completeness of the dataset with respect to NNP and fine colloids in multiple water compartments of a single forest watershed this study adds to the hitherto underexplored role of NNP and fine colloids in natural forest watersheds., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Liposomal formulations for the treatment of cancer and other diseases are the most common form of nanotechnology enabled pharmaceuticals (NEPs) submitted for market approval and in clinical application today. The accurate characterization of their physical-chemical properties is a key requirement; in particular, size, size distribution, shape, and physical-chemical stability are key among properties that regulators identify as critical quality attributes. Here we develop and validate an optimized method, based on multi-detector asymmetrical-flow field flow fractionation (MD-AF4) to accurately and reproducibly separate liposomal drug formulations into their component populations and to characterize their associated size and size distribution, whether monomodal or polymodal in nature. In addition, the results show that the method is suitable to measure liposomes in the presence of serum proteins and can yield information on the shape and physical stability of the structures. The optimized MD-AF4 based method has been validated across different instrument platforms, three laboratories, and multiple drug formulations following a comprehensive analysis of factors that influence the fractionation process and subsequent physical characterization. Interlaboratory reproducibility and intra-laboratory precision were evaluated, identifying sources of bias and establishing criteria for the acceptance of results. This method meets a documented high priority need in regulatory science as applied to NEPs such as Doxil and can be adapted to the measurement of other NEP forms (such as lipid nanoparticle therapeutics) with some modifications. Overall, this method will help speed up development of NEPS, and facilitate their regulatory review, ultimately leading to faster translation of innovative concepts from the bench to the clinic. Additionally, the approach used in this work (based on international collaboration between leading non-regulatory institutions) can be replicated to address other identified gaps in the analytical characterization of various classes of NEPs. Finally, a plan exists to pursue more extended interlaboratory validation studies to advance this method to a consensus international standard., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Regular algal blooms are occurring in Taihu lake, which may be triggered by resuspension of sediments containing relevant amounts of phosphorus. Therefore, our study aims at quantification of phosphorus concentrations bound to suspended particulate matter in Taihu water samples to investigate this hypothesis. A field flow fractionation (FFF) method online with ICP-MS detection was developed to achieve an overview on particulate fractions of phosphorus and related elements including Fe, Al and C from the low nanometer to the low micrometer size range. Mass balance of dissolved and particulate elemental contents was established for quality control purpose and indicated low recovery of Fe, Al and P. Complementary determination of volume based particle size distribution by dynamic imaging analysis showed a majority of particle volume and thus mass in particles with size >5 μm. In order to address this challenge, the upper particle size limit of FFF online with ICP-MS was for the first time investigated in detail using well characterised monodisperse latex particles as model for organic matter in the low micrometer size range including microalgae. The effect of pre-filtration of the sample as well as the contribution of sample introduction via three different interfaces including micromist nebuliser/spray chamber, direct injection nebulisation and APEX with heated spray chamber and solvent removal by condensation on the particulate carbon recovery was studied by ICP-MS detection. The same instrumental setup was also applied for the characterisation of particulate elemental contents in the Taihu water samples as far as possible. Significant improvement of the detected particulate fraction in Taihu water samples was achieved by increasing the membrane pore size for pre-filtration and by using the APEX for introduction of the eluate from FFF into ICP-MS., (Copyright © 2019 Elsevier B.V. All rights reserved.)

This study is about linking preparative processes of nanoparticles with the morphology of the nanoparticles and with their efficiency in delivering payloads intracellularly. The nanoparticles are composed of hyaluronic acid (HA) and chitosan; the former can address a nanoparticle to cell surface receptors such as CD44, the second allows both for entrapment of nucleic acids and for an endosomolytic activity that facilitates their liberation in the cytoplasm. Here, we have systematically compared nanoparticles prepared either A) through a two-step process based on intermediate (template) particles produced via ionotropic gelation of chitosan with triphosphate (TPP), which are then incubated with HA, or B) through direct polyelectrolyte complexation of chitosan and HA. Here we demonstrate that HA is capable to quantitatively replace TPP in the template process and significant aggregation takes place during the TPP-HA exchange. The templated chitosan/HA nanoparticles therefore have a mildly larger size (measured by dynamic light scattering alone or by field flow fractionation coupled to static or dynamic light scattering), and above all a higher aspect ratio ( R g / R H ) and a lower fractal dimension. We then compared the kinetics of uptake and the (antiluciferase) siRNA delivery performance in murine RAW 264.7 macrophages and in human HCT-116 colorectal tumor cells. The preparative method (and therefore the internal particle morphology) had little effect on the uptake kinetics and no statistically relevant influence on silencing (templated particles often showing a lower silencing). Cell-specific factors, on the contrary, overwhelmingly determined the efficacy of the carriers, with, e.g., those containing low-MW chitosan performing better in macrophages and those with high-MW chitosan in HCT-116., (Copyright © 2019, Gennari et al.; licensee Beilstein-Institut.)

Characterization of natural nanomaterial (NNM) physicochemical properties - such as size, size distribution, elemental composition and elemental ratios - is often hindered by lack of methods to disperse NNMs from environmental samples. This study evaluates the effect of extractant composition, pH, and ionic strength on soil NNM extraction in term of recovery and release of primary particles/small aggregate sizes (i.e., <200 nm). The extracted NNMs were characterized for hydrodynamic diameter and zeta potential by dynamic light scattering and laser Doppler electrophoresis, natural organic matter desorption by UV-Vis spectroscopy, element composition by inductively coupled plasma-mass spectroscopy (ICP-MS), size based elemental distribution by field flow fractionation coupled to ICP-MS, and morphology by transmission electron microscopy. The extracted NNM concentrations increased following the order of NaOH ≤ Na 2 CO 3  < Na 2 C 2 O 4  < Na 4 P 2 O 7 . Na 4 P 2 O 7 was the most efficient extractant and results in 2-12 folds higher NNM extraction than other extractants. The Na 4 P 2 O 7 extracted NNMs exhibited narrower size distribution with smaller modal size relative to NaOH, Na 2 CO 3 , Na 2 C 2 O 4 extracted NNMs. Thus, Na 4 P 2 O 7 enhances the extraction of primary NNMs and/or smaller NNM aggregates (i.e., size <200 nm). Na 4 P 2 O 7 promote soil microaggregates breakup and release of NNMs by reducing free multivalent cation concentration in soil pore water by forming metal-phosphate complexes and by enhancing NNM surface charge via phosphate sorption on NNM surfaces. Additionally, the extracted NNM concentrations increased with the increase in extractant concentration and pH, except at 100 mM where the high ionic strength might have induced NNM aggregation. The improved NNM-extraction will improve the overall understanding of the physicochemical properties of NNMs in environmental systems. This study presents the key properties of NNMs that can be used as background information to differentiate engineered nanomaterials (ENMs) from NNMs in complex environmental media., (Copyright © 2019 Elsevier B.V. All rights reserved.)

This paper describes the development of an integrated system using a dry film resistant (DFR) microfluidic channel consisting of pulsed field dielectrophoretic field-flow-fractionation (DEP-FFF) separation and optical detection. The prototype chip employs the pulse DEP-FFF concept to separate the cells ( Escherichia coli and Saccharomyces cerevisiae ) from a continuous flow, and the rate of release of the cells was measured. The separation experiments were conducted by changing the pulsing time over a pulsing time range of 2⁻24 s and a flow rate range of 1.2⁻9.6 μ L min - 1 . The frequency and voltage were set to a constant value of 1 M Hz and 14 V pk-pk, respectively. After cell sorting, the particles pass the optical fibre, and the incident light is scattered (or absorbed), thus, reducing the intensity of the transmitted light. The change in light level is measured by a spectrophotometer and recorded as an absorbance spectrum. The results revealed that, generally, the flow rate and pulsing time influenced the separation of E. coli and S. cerevisiae . It was found that E. coli had the highest rate of release, followed by S. cerevisiae . In this investigation, the developed integrated chip-in-a lab has enabled two microorganisms of different cell dielectric properties and particle size to be separated and subsequently detected using unique optical properties. Optimum separation between these two microorganisms could be obtained using a longer pulsing time of 12 s and a faster flow rate of 9.6 μ L min - 1 at a constant frequency, voltage, and a low conductivity.

Both fully (98%) and partially (88%) hydrolyzed poly(vinyl alcohol) (PVA) are characterized by aqueous gel permeation chromatography (GPC) coupled with multi-angle laser light scattering. Polymer adsorption on hydrophilic GPC columns is observed for partially hydrolyzed PVA but not for fully hydrolyzed PVA, which is driven by the hydrophobic interactions between the residual acetate groups on PVA and the stationary phase. Desorption of these partially hydrolyzed PVA chains from the packing materials is monitored by decay of the light scattering signal. It is found that the desorption of these polymer chains from porous particles is non-exponential and follows stretched exponential kinetics with index 0.36. It is also shown that asymmetric flow field flow fractionation can be successfully used for the molar mass characterization of PVA, irrespective of the degree of hydrolysis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

An acoustic field flow fractionation (FFF) device was developed to fractionate a micro-particle mixture on the basis of the particle diameter using an acoustic force field in a carrier liquid flow. In the acoustic FFF channel used in the device, ultrasound waves generated from piezoelectric transducers driven by a sinusoidal signal of 2.02 Mhz propagated into the carrier liquid flow and built up a quarter-wavelength ultrasound standing wave field across the channel height. It was experimentally demonstrated that the acoustic field with a pressure node plane at the bottom surface of the channel reduced the thickness of the particle diffusion layer in a stagnant liquid proportional to the applied voltage driving the piezoelectric transducer. In the size-dependent particle separation, the particle mixture flowing through the acoustic FFF channel experienced an acoustic radiation force in the gravitational direction. As a result, suppressing the diffusion of small particles, particles were transported along the bottom surface of the channel with the local velocity of the carrier liquid at the particle center. The developed acoustic FFF device successfully fractionated a fluorescent micro-particle mixture (1, 3, 5, and 10 μm diameter), whereas the 3 and 5 μm particles were not fractionated in the FFF device using only the gravitational force field due to the diffusion of 3 μm particles., (Copyright © 2018 Elsevier B.V. All rights reserved.)

For best photosensitizer activity phthalocyanine dyes used in photodynamic therapy should be molecularly dispersed. Polyethylene glycol-block-polylactide derivatives presenting benzyl side-groups were synthesized to encapsulate a highly lipophilic phthalocyanine dye (AlClPc) and evaluate the effect of π-π interactions on the nanocarrier colloidal stability and dye dispersion. Copolymers with 0, 1, 2 and 6 mol% of benzyl glycidyl ether (BGE) were obtained via polyethylene glycol initiated ring-opening copolymerization of D,l-lactide with BGE. The block copolymers formed stable, monodisperse nanospheres with low in vitro cytotoxicity. AlClPc loading increased the nanosphere size and affected their colloidal stability. The photo-physical properties of the encapsulated dye, studied in batch and after separation by field flow fractionation, demonstrated the superiority of plain PEG-PLA over BGE-containing copolymers in maintaining the dye in its monomeric (non-aggregated) form in aqueous suspension. High dye encapsulation and sustained dye release suggest that these nanocarriers are good candidates for photodynamic therapy., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Applications of asymmetrical flow field-flow fractionation (AF4) continue to expand rapidly in the fields of nanotechnology and biotechnology. In particular, AF4 has proven valuable for the separation and analysis of particles, biomolecular species (e.g., proteins, bacteria) and polymers (natural and synthetic), ranging in size from a few nanometers to several micrometers. The separation of non-spheroidal structures (e.g., rods, tubes, etc.) with primary dimensions in the nanometer regime, is a particularly challenging application deserving of greater study and consideration. The goal of the present study was to advance current understanding of the mechanism of separation of rod-like nano-objects in the AF4 channel. To achieve this, we have systematically investigated a series of commercially available cetyltrimethylammonium bromide stabilized gold nanorods (AuNRs), with aspect ratios from 1.7 to 10. Results show clearly that the retention time is principally dependent on the translational diffusion coefficient of the AuNRs. Equations used to calculate translational and rotational diffusion coefficients (cylinder and prolate ellipsoid models) yield similarly good fits to experimental data. Well characterized gold nanorods (length and diameter by transmission electron microscopy) can be used as calibrants for AF4 measurements allowing one to determine the aspect ratio of nanorod samples based on their retention times. Graphical abstract ᅟ.

The interaction of polymer nanocapsules (NC) prepared from four biodegradable polyesters with variable polymer hydrophobicity (PCL, PLA, PLGA and PLA-PEG) was investigated in the non-phagocytic Vero, Caco-2 and HepG2 cell lines. The NC, labeled with the highly lipophilic fluorescent indocarbocyanine dye DIL, had very similar sizes (approx. 140 nm) and negative zeta-potentials. Asymmetric flow field-flow fractionation evidenced NC colloidal stability and negligible transfer of the dye to serum proteins in the incubation medium. The cytotoxicity of the NC was evaluated via MTT assay over a large polymer concentration range (1-1000 μg/mL) and time of exposure (2, 24 and 48 h). The NC were safe in vitro up to a concentration of approx. 100 μg/mL or higher, depending on the cell line and nature of the polymer. Vero cells were more sensitive to the NC, in particular NC of the more hydrophobic polymer. The cells were exposed to endocytosis inhibitors, incubated with NC, and the cell-associated fluorescence was quantified by spectrofluorometry. HepG2 cells presented a 1.5-2-fold higher endocytic capacity than Caco-2 and Vero cells. The main mechanism of NC uptake was caveolin-mediated endocytosis in HepG2 and Vero cells, and macropinocytosis in Caco-2 cells. Polymer hydrophobicity had an effect on the level of NC associated to HepG2 cells and to a lesser extent on the endocytosis mechanisms in Vero and Caco-2 cells. The NC uptake levels and endocytosis mechanisms differed significantly between cell lines tested., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Basic and applied virus research requires specimens that are purified to high homogeneity. Thus, there is much interest in the efficient production and purification of viruses and their subassemblies. Advances in the production steps have shifted the bottle neck of the process to the purification. Nonetheless, the development of purification techniques for different viruses is challenging due to the complex biological nature of the infected cell cultures as well as the biophysical and -chemical differences in the virus particles. We used bacteriophage ϕ6 as a model virus in our attempts to provide a new purification method for enveloped viruses. We compared asymmetrical flow field-flow fractionation (AF4)-based virus purification method to the well-established ultracentrifugation-based purification of ϕ6. In addition, binding of ϕ6 virions to monolithic anion exchange columns was tested to evaluate their applicability in concentrating the AF4 purified specimens. Our results show that AF4 enables one-hour purification of infectious enveloped viruses with specific infectivity of ~1 × 10 13  PFU/mg of protein and ~65-95% yields. Obtained purity was comparable with that obtained using ultracentrifugation, but the yields from AF4 purification were 2-3-fold higher. Importantly, high quality virus preparations could be obtained directly from crude cell lysates. Furthermore, when used in combination with in-line light scattering detectors, AF4 purification could be coupled to simultaneous quality control of obtained virus specimen., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The use of nanotechnology in medical products has been demonstrated at laboratory scale, and many resulting nanomedicines are in the translational phase toward clinical applications, with global market trends indicating strong growth of the sector in the coming years. The translation of nanomedicines toward the clinic and subsequent commercialization may require the development of new or adaptation of existing standards to ensure the quality, safety and efficacy of such products. This work addresses some identified needs, and illustrates the shortcomings of currently used standardized methods when applied to medical-nanoparticles to assess particle size, drug loading, drug release and in vitro safety. Alternative physicochemical, and in vitro toxicology methods, with the potential to qualify as future standards supporting the evaluation of nanomedicine are provided.

Silver nanoparticles (Ag-NP) are used in various consumer products and are one of the most prevalent metallic nanoparticle in commodities and are released into the environment. Transport behavior of Ag-NP in groundwater is one important aspect for the assessment of environmental impact and protection of drinking water resources in particular. Ag-NP transport processes in saturated single-fractured sandstones using triaxial flow cell experiments with different kind of sandstones is investigated. Ag-NP concentration and size are analyzed using flow field-flow fractionation and coupled SEM-EDX analysis. Results indicate that Ag-NP are more mobile and show generally lower attachment on rock surface compared to experiments in undisturbed sandstone matrix and partially fractured sandstones. Ag-NP transport is controlled by the characteristics of matrix porosity, time depending blocking of attachment sites and solute chemistry. Where Ag-NP attachment occur, it is heterogeneously distributed on the fracture surface., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The impact and behavior of engineered nanomaterials (ENMs) entering the environment is an important issue due to their growing use in consumer and agricultural products. Their mobility and fate in the environment are heavily impacted by their interactions with natural particle components of saturated sediments and soils. In this study, functionalized gold nanoparticles (AuNPs - used as model ENMs) were spiked into complex solid-containing media (standard soils and estuarine sediment in moderately hard water). AuNPs were characterized in the colloidal extract (< 1 μm) following centrifugal separation of the non-colloidal phase, using different analytical techniques including asymmetric-flow field-flow fractionation and single particle inductively coupled plasma mass spectrometry. Attachment of functionalized AuNPs to the soil particles did not significantly depend on their concentration or surface coating (citrate, bPEI, PVP, PEG). Similarly, UV degradation of coatings did not substantially alter their recovery. Conversely, the presence of natural organic matter (NOM) is a key factor in their adhesion to matrix particles, by decreasing the predicted influence of native surface chemistry and functional coatings. A kinetic experiment performed over 48 h showed that attachment to soil colloids is rapid and that hetero-aggregation is dominant. These results suggest that transport of ENMs away from the point of discharge (or entry) could be limited in soils and sediments, but additional experiments under more realistic and dynamic field conditions would be necessary to confirm this more generally. Transport properties may also differ substantially in matrices where NOM is largely absent or otherwise sequestered or when dissolution of ENMs is an important factor.

In the last 10 years, asymmetrical flow field flow fractionation (AF4) has been one of the most promising approaches to characterize colloidal particles. Nevertheless, despite its potentialities, it is still considered a complex technique to set up, and the theory is difficult to apply for the characterization of complex samples containing submicron particles and nanoparticles. In the present work, we developed and propose a simple analytical strategy to rapidly determine the presence of several submicron populations in an unknown sample with one programmed AF4 method. To illustrate this method, we analyzed polystyrene particles and fullerene aggregates of size covering the whole colloidal size distribution. A global and fast AF4 method (method O) allowed us to screen the presence of particles with size ranging from 1 to 800 nm. By examination of the fractionating power F d , as proposed in the literature, convenient fractionation resolution was obtained for size ranging from 10 to 400 nm. The global F d values, as well as the steric inversion diameter, for the whole colloidal size distribution correspond to the predicted values obtained by model studies. On the basis of this method and without the channel components or mobile phase composition being changed, four isocratic subfraction methods were performed to achieve further high-resolution separation as a function of different size classes: 10-100 nm, 100-200 nm, 200-450 nm, and 450-800 nm in diameter. Finally, all the methods developed were applied in characterization of nanoplastics, which has received great attention in recent years. Graphical Absract Characterization of the nanoplastics by asymmetrical flow field flow fractionation within the colloidal size range.

The effect of polymeric nanocapsule dose on plasmatic and liver concentrations 20min after intravenous administration in mice was evaluated. Nanocapsules were prepared with different polymers, namely, poly(D,L-lactide) (PLA), polyethylene glycol-block-poly(D,L-lactide) (PLA-PEG), and PLA with chitosan (PLA-Cs) and compared with a nanoemulsion. These formulations were labelled with a phthalocyanine dye for fluorescent detection. The nanostructures had narrow size distributions upon separation by asymmetric flow field flow fractionation with static and dynamic light scattering detection, with average hydrodynamic diameters in the 130-300nm range, negative zeta potentials, except PLA-Cs nanocapsules, which had a positive zeta potential. Flow cytometry revealed uptake mostly by monocytes and neutrophils in mice and human blood. PLA nanocapsules and the nanoemulsion showed dose-dependent plasma concentrations, where the percentage of plasmatic fluorescence increased with increasing administered dose. In contrast, PLA-PEG nanocapsules led to a dose-independent plasmatic profile. PLA-Cs nanocapsules showed the lowest plasmatic and liver levels of fluorescence at all administered doses and significant intravenous toxicity in mice. This work demonstrates the importance of considering the nanocarrier dose when evaluating pharmacokinetic and biodistribution data and emphasizes the role of surface features in determining the plasmatic and liver concentrations of a poorly soluble lipophilic encapsulated compound., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Field flow fractionation (FFF) techniques are used to successfully characterize several nanomaterials by sizing nano-entities and producing information about the aggregation/agglomeration state of nanoparticles. By coupling FFF techniques to specific detectors, researchers can determine particle-size distributions (PSDs), expressed as mass-based or number-based PSDs. This review considers FFF applications in the food, biomedical, and environmental sectors, mostly drawn from the past 4 y. It thus underlines the prominent role of asymmetrical flow FFF within the FFF family. By concisely comparing FFF techniques with other techniques suitable for sizing nano-objects, the advantages and the disadvantages of these instruments become clear. A consideration of select recent publications illustrates the state of the art of some lesser-known FFF techniques and innovative instrumental set-ups.

Cell specific ligand molecules are attached to polyethylene glycol (PEG) modified nanoparticles to enhance the drug delivery efficiency. The tethered PEG would interfere in ligand recognition as well as providing biocompatibility to the nanoparticles. The denser PEG can give the greater biocompatibility, while should more hamper the ligand recognition. Therefor it is important to tune PEG density at an appropriate amount to compensate these two factors. In this study, we prepared a series of nanoparticles composed of α-mannose-bearing lipid, dioleoyltrimethyl ammoniumpropane (DOTAP) and 1,2-distearoyl-sn-glycero -3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (DSPE-PEG2000). The nanoparticles were characterized with dynamic light scattering (DLS), field flow fractionation (FFF), small angle X-ray scattering (SAXS), and quartz-crystal microbalance (QCM). Based on the structural parameter obtained from DLS, SAXS, and FFF, we determined the PEG crowding parameter (σ) quantitatively; when σ=1.0, the PEG chain occupies the same value on the surface as when the chain is present in the unperturbed state, and at σ=1.0, the PEG chains start to contact each other. We found that the recognition ability had the maximum around σ∼0.75 and there was the critical composition at σ=1.0 for which the recognition was drastically reduced. The present results demonstrated that quantitative characterization and controlling the PEG density are key to designing effective targeting delivery system., (Copyright © 2016. Published by Elsevier B.V.)

We demonstrate the use of inverse supercritical carbon dioxide (scCO2) extraction as a novel method of sample preparation for the analysis of complex nanoparticle-containing samples, in our case a model sunscreen agent with titanium dioxide nanoparticles. The sample was prepared for analysis in a simplified process using a lab scale supercritical fluid extraction system. The residual material was easily dispersed in an aqueous solution and analyzed by Asymmetrical Flow Field-Flow Fractionation (AF4) hyphenated with UV- and Multi-Angle Light Scattering detection. The obtained results allowed an unambiguous determination of the presence of nanoparticles within the sample, with almost no background from the matrix itself, and showed that the size distribution of the nanoparticles is essentially maintained. These results are especially relevant in view of recently introduced regulatory requirements concerning the labeling of nanoparticle-containing products. The novel sample preparation method is potentially applicable to commercial sunscreens or other emulsion-based cosmetic products and has important ecological advantages over currently used sample preparation techniques involving organic solvents., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The control of gold nanorod (GNR) solution-based syntheses has been hindered in part by the inability to examine and control the conversion of precursor seed populations to anisotropic materials, which have resulted in low yields of desired products and limited their commercial viability. The advantages offered by tandem separation and characterization methods utilizing asymmetric-flow field flow fractionation (A4F) are principally achieved as a result of their non-disruptive nature (minimizing artefacts), fast throughput, and in-situ analysis. With hyphenated A4F methods, resolved populations of seeds and secondary products, up to long aspect ratio rods, have been achieved and exemplify progress towards elucidating mechanistic aspects of formation and thus rational design. While there have been previously reported studies on A4F separation of GNRs, to our knowledge, this is the first published investigation of in situ GNR growth, separation, and characterization based on A4F, where its utilization in this capacity goes beyond traditional separation analysis. By using hydroquinone as the reducing agent, the conversion of the initial seed population to a distribution of products, including the GNRs, could be monitored in real time using A4F hyphenated with a diode array detector. Transmission electron microscopy confirms that the number of peaks observed during fractionation corresponds with size and shape dispersity. This proof-of-principle study introduces A4F as a technique that establishes a foundation for future mechanistic studies on the growth of GNRs from gold seeds, including conversion of the seed population to initial products, a topic highly relevant to advancing progress in nanomanufacturing.

Asymmetrical Flow Field-Flow Fractionation (AF4) is a separation technique applicable to particles over a wide size range. Despite the many advantages of AF4, its adoption in routine particle analysis is somewhat limited by the large footprint of currently available separation cartridges, extended analysis times and significant solvent consumption. To address these issues, we describe the fabrication and characterization of miniaturized AF4 cartridges. Key features of the down-scaled platform include simplified cartridge and reagent handling, reduced analysis costs and higher throughput capacities. The separation performance of the miniaturized cartridge is assessed using certified gold and silver nanoparticle standards. Analysis of gold nanoparticle populations indicates shorter analysis times and increased sensitivity compared to conventional AF4 separation schemes. Moreover, nanoparticulate titanium dioxide populations exhibiting broad size distributions are analyzed in a rapid and efficient manner. Finally, the repeatability and reproducibility of the miniaturized platform are investigated with respect to analysis time and separation efficiency.

In order to assess the environmental risks of a compound it is imperative to have suitable and reliable techniques for its determination in environmental matrices. In this paper, we focused on a method development for the recently introduced online coupling of a field flow fractionation (FFF) system to an Orbitrap-HRMS, that allows the simultaneous size and concentration determination of different aqueous fullerene aggregates and their concentrations in different size fractions. A 0.05% NH4OH solution in water was identified as the best carrier liquid for the analysis of the three different aqueous fullerene suspensions (C60 [60], [6,6]-phenyl-C61 butyric acid methyl ester ([60]PCBM) and [6,6]-(bis)phenyl-C61 butyric acid methyl ester ([60]bisPCBM)). The multi-angle light scattering (MALS) data received after employing the ammonia solution was consistent with both the theory and calibration using well defined Au and latex particles. The LODs obtained using Orbitrap HRMS detection were 0.1 μg L(-1) for an injection volume of 100 μL which are significantly better than the LODs obtained by using UV (20 μg L(-1)) and MALS detectors (5 μg L(-1)). However, these LODs can be further improved as in theory there is no limit to the amount of sample that can be injected into the FFF. Environmental samples (river and sewage water) were spiked with fullerenes and the fractograms obtained for these samples revealed that the matrix does affect the size of fullerene aggregates. Information on the size distribution can be useful for the risk assessment of these particles., (Copyright © 2015 Elsevier B.V. All rights reserved.)

In situ measurements of labile metal species using diffusive gradients in thin films (DGT) passive samplers are based on the diffusion rates of individual species. Although most studies have dealt with chemically isolated humic substances, the diffusion of dissolved organic matter (DOM) across the hydrogel is not well understood. In this study, the diffusion coefficient (D) and molecular weight (MW) of 11 aquatic DOM and 4 humic substances (HS) were determined. Natural, unaltered aquatic DOM was capable of diffusing across the diffusive gel membrane with D values ranging from 2.48×10(-6) to 5.31×10(-6) cm(2) s(-1). Humic substances had diffusion coefficient values ranging from 3.48×10(-6) to 6.05×10(-6) cm(2) s(-1), congruent with previous studies. Molecular weight of aquatic DOM and HS samples (∼500-1750 Da) measured using asymmetrical flow field-flow fractionation (AF4) strongly influenced D, with larger molecular weight DOM having lower D values. No noticeable changes in DOM size properties were observed during the diffusion process, suggesting that DOM remains intact following diffusion across the diffusive gel. The influence of molecular weight on DOM mobility will assist in further understanding and development of the DGT technique and the uptake and mobility of contaminants associated with DOM in aquatic environments., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Compared to other sub-techniques of field flow fractionation (FFF), cyclical electrical field flow fractionation (CyElFFF) is a relatively new method with many opportunities remaining for improvement. One of the most important limitations of this method is the separation of particles smaller than 100nm. For such small particles, the diffusion rate becomes very high, resulting in severe reductions in the CyElFFF separation efficiency. To address this limitation, we modified the electrical circuitry of the ElFFF system. In all earlier ElFFF reports, electrical power sources have been directly connected to the ElFFF channel electrodes, and no alteration has been made in the electrical circuitry of the system. In this work, by using discrete electrical components, such as resistors and diodes, we improved the effective electric field in the system to allow high resolution separations. By modifying the electrical circuitry of the ElFFF system, high resolution separations of 15 and 40nm gold nanoparticles were achieved. The effects of applying different frequencies, amplitudes and voltage shapes have been investigated and analyzed through experiments., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Nanoparticles (NPs) adsorb proteins when in the biological matrix, and the resulted protein corona could affect NP-cell interactions. The corona has a dynamic nature with the adsorbed proteins constantly exchanging with the free proteins in the matrix at various rates. The rapidly exchanging proteins compose the soft corona, which responds more dynamically to environment changes than the hard corona established by the ones with slow exchange rates. In the present study, the corona formed on the superparamagnetic iron oxide NPs (SPIONs) in human serum was studied by flow field-flow fractionation and ultracentrifugation, which rapidly differentiated the corona proteins based on their exchange rates. By varying the surface hydrophobicity of the SPIONs with a core size around 10 nm, we found out that, the more hydrophobic surface ligand attracted proteins with higher surface hydrophobicity and formed a more dynamic corona with a larger portion of the involved proteins with fast exchange rates. Increasing the core diameter of the SPIONs but keeping the surface ligand the same could also result in a more dynamic corona. A brief investigation of the effect on the cellular uptake of SPIONs using one selected corona protein, transferrin, was conducted. The result showed that, only the stably bound transferrin could significantly enhance cellular uptake, while transferrin bound in a dynamic nature had negligible impact. Our study has led to a better understanding of the relationship between the particle properties and the dynamic nature of the corona, which can help with design of nanomaterials with higher biocompatibility and higher efficacy in biosystems for biomedical applications.

Transport behavior and fate of engineered silver nanoparticles (AgNP) in the subsurface is of major interest concerning soil and groundwater protection in order to avoid groundwater contamination of vital resources. Sandstone aquifers are important groundwater resources which are frequently used for public water supply in many regions of the world. The objective of this study is to get a better understanding of AgNP transport behavior in partially fractured sandstones. We executed AgNP transport studies on partially fissured sandstone drilling cores in laboratory experiments. The AgNP concentration and AgNP size in the effluent were analyzed using flow field-flow fractionation mainly. We employed inverse mathematical models on the measured AgNP breakthrough curves to identify and quantify relevant transport processes. Physicochemical filtration, time-dependent blocking due to filling of favorable attachment sites and colloid-facilitated transport were identified as the major processes for AgNP mobility. Physicochemical filtration was found to depend on solute chemistry, mineralogy, pore size distribution and probably on physical and chemical heterogeneity. Compared to AgNP transport in undisturbed sandstone matrix reported in the literature, their mobility in partially fissured sandstone is enhanced probably due to larger void spaces and higher hydraulic conductivity., (Copyright © 2014 Elsevier B.V. All rights reserved.)

This tutorial proposes a comprehensive and rational measurement strategy that provides specific guidance for the application of asymmetric-flow field flow fractionation (A4F) to the size-dependent separation and characterization of nanoscale particles (NPs) dispersed in aqueous media. A range of fractionation conditions are considered, and challenging applications, including industrially relevant materials (e.g., metal NPs, asymmetric NPs), are utilized in order to validate and illustrate this approach. We demonstrate that optimization is material dependent and that polystyrene NPs, widely used as a reference standard for retention calibration in A4F, in fact represent a class of materials with unique selectivity, recovery and optimal conditions for fractionation; thus use of these standards to calibrate retention for other materials must be validated a posteriori. We discuss the use and relevance of different detection modalities that can potentially yield multi-dimensional and complementary information on NP systems. We illustrate the fractionation of atomically precise nanoclusters, which are the lower limit of the nanoscale regime. Conversely, we address the upper size limit for normal mode elution in A4F. The protocol for A4F fractionation, including the methods described in the present work is proposed as a standardized strategy to realize interlaboratory comparability and to facilitate the selection and validation of material-specific measurement parameters and conditions. It is intended for both novice and advanced users of this measurement technology., (Published by Elsevier B.V.)

In the field of nanomedicine, cationic polymers are the subject of intensive research and represent promising carriers for genetic material. The detailed characterization of these carriers is essential since the efficiency of gene delivery strongly depends on the properties of the used polymer. Common characterization methods such as size exclusion chromatography (SEC) or mass spectrometry (MS) suffer from problems, e.g. missing standards, or even failed for cationic polymers. As an alternative, asymmetrical flow field-flow fractionation (AF4) was investigated. Additionally, analytical ultracentrifugation (AUC) and (1)H NMR spectroscopy, as well-established techniques, were applied to evaluate the results obtained by AF4. In this study, different polymers of molar masses between 10 and 120kgmol(-1) with varying amine functionalities in the side chain or in the polymer backbone were investigated. To this end, some of the most successful gene delivery agents, namely linear poly(ethylene imine) (LPEI) (only secondary amines in the backbone), branched poly(ethylene imine) (B-PEI) (secondary and tertiary amino groups in the backbone, primary amine end groups), and poly(l-lysine) (amide backbone and primary amine side chains), were characterized. Moreover, poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA), poly(2-(amino)ethyl methacrylate) (PAEMA), and poly(2-(tert-butylamino)ethyl methacrylate) (PtBAEMA) as polymers with primary, secondary, and tertiary amines in the side chain, have been investigated. Reliable results were obtained for all investigated polymers by AF4. In addition, important factors for all methods were evaluated, e.g. the influence of different elution buffers and AF4 membranes. Besides this, the correct determination of the partial specific volume and the suppression of the polyelectrolyte effect are the most critical issues for AUC investigations., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Recent advances in instrumentation and data analysis in field flow fractionation and multi-angle light scattering (FFF-MALS) have enabled greater use of this technique to characterize and quantitate viruses. In this study, the FFF-MALS technique was applied to the characterization and quantitation of type A influenza virus particles to assess its usefulness for vaccine preparation. The use of FFF-MALS for quantitation and measurement of control particles provided data accurate to within 5% of known values, reproducible with a coefficient of variation of 1.9%. The methods, sensitivity and limit of detection were established by analyzing different volumes of purified virus, which produced a linear regression with fitting value R2 of 0.99. FFF-MALS was further applied to detect and quantitate influenza virus in the supernatant of infected MDCK cells and allantoic fluids of infected eggs. FFF fractograms of the virus present in these different fluids revealed similar distribution of monomeric and oligomeric virions. However, the monomer fraction of cell grown virus had greater size variety. Notably, β-propialactone (BPL) inactivation of influenza viruses did not influence any of the FFF-MALS measurements. Quantitation analysis by FFF-MALS was compared to infectivity assays and real-time RT-PCR (qRT-PCR) and the limitations of each assay were discussed., (Published by Elsevier B.V.)

We present numerical simulation of separating magnetic particles with different magnetic susceptibilities by magnetic chromatography using a high-temperature superconducting bulk magnet. The transient transport is numerically simulated for two kinds of particles having different magnetic susceptibilities. The time evolutions were calculated for the particle concentration in the narrow channel of the spiral arrangement placed in the magnetic field. The field is produced by the highly magnetized high-temperature superconducting bulk magnet. The numerical results show the flow velocity difference of the particle transport corresponding to the difference in the magnetic susceptibility, as well as the possible separation of paramagnetic particles of 20 nm diameter.