Multi-environment and multi-parameter screening of stability and coating efficiency of gold nanoparticle bioconjugates in application media.

Gold nanoparticles (AuNPs) and their biocompatible conjugates find wide use as transducers in (bio)sensors and as Nano-pharmaceutics. The study of the interaction between AuNPs and proteins in representative application media helps to better understand their intrinsic behaviors. A multi-environment, multi-parameter screening strategy is proposed based on asymmetric flow field flow fractionation (AF4)-multidetector.

Emerging technologies for quality control of cell-based, advanced therapy medicinal products.

Advanced therapy medicinal products (ATMP) are complex medicines based on gene therapy, somatic cell therapy, and tissue engineering. These products are rapidly arising as novel and promising therapies for a wide range of different clinical applications. The process for the development of well-established ATMPs is challenging.

Comprehensive analysis of colloid formation, distribution, and properties of monovarietal red wines using asymmetrical flow field-flow fractionation with online multidetection.

Red wine colloids, crucial in determining wine quality and stability, are understudied due to inadequate techniques for studying them effectively in the natural wine environment. Recently, Asymmetrical Flow Field-flow Fractionation (AF4) with online multidetection has emerged as a novel analytical tool for quantifying, fractionating, and characterizing red wine colloids in their native state. This study aimed to characterize the colloidal composition of 24 monovarietal Italian wines produced without filtration, oak contact, fining treatments, malolactic fermentation, macerating enzymes or ageing on yeast lees.

Native characterization and QC profiling of human amniotic mesenchymal stromal cell vesicular fractions for secretome-based therapy.

Human amniotic mesenchymal stromal cells (hAMSCs) have unique immunomodulatory properties making them attractive candidates for regenerative applications in inflammatory diseases. Most of their beneficial properties are mediated through their secretome. The bioactive factors concurring to its therapeutic activity are still unknown.

Emerging Microfluidic Tools for Simultaneous Exosomes and Cargo Biosensing in Liquid Biopsy: New Integrated Miniaturized FFF-Assisted Approach for Colon Cancer Diagnosis.

The early-stage diagnosis of cancer is a crucial clinical need. The inadequacies of surgery tissue biopsy have prompted a transition to a less invasive profiling of molecular biomarkers from biofluids, known as liquid biopsy. Exosomes are phospholipid bilayer vesicles present in many biofluids with a biologically active cargo, being responsible for cell-to-cell communication in biological systems.

The dual nature of biomimetic melanin.

Melanin-inspired nanomaterials offer unique photophysical, electronic and radical scavenging properties that are widely explored for health and environmental preservation, or energy conversion and storage. The incorporation of functional melanin building blocks in more complex nanostructures or surfaces is typically achieved a bottom-up approach starting from a molecular precursor, in most cases dopamine. Here we demonstrate that indeed, the oxidative polymerization of dopamine, for the synthesis of melanin-like polydopamine (PDA), leads to the simultaneous formation of more than one nanosized species with different compositions, morphologies and properties.

Liposomes characterization for market approval as pharmaceutical products: Analytical methods, guidelines and standardized protocols.

Liposomes are nano-sized lipid-based vesicles widely studied for their drug delivery capabilities. Compared to standard carries they exhibit better properties such as improved site-targeting and drug release, protection of drugs from degradation and clearance, and lower toxic side effects. At present, scientific literature is rich of studies regarding liposomes-based systems, while 14 types of liposomal products have been authorized to the market by EMA and FDA and many others have been approved by national agencies.

Field-Flow Fractionation in Molecular Biology and Biotechnology.

Field-flow fractionation (FFF) is a family of single-phase separative techniques exploited to gently separate and characterize nano- and microsystems in suspension. These techniques cover an extremely wide dynamic range and are able to separate analytes in an interval between a few nm to 100 µm size-wise (over 15 orders of magnitude mass-wise). They are flexible in terms of mobile phase and can separate the analytes in native conditions, preserving their original structures/properties as much as possible.

Native Study of the Behaviour of Magnetite Nanoparticles for Hyperthermia Treatment during the Initial Moments of Intravenous Administration.

Magnetic nanoparticles (MNPs) present outstanding properties making them suitable as therapeutic agents for hyperthermia treatments. Since the main safety concerns of MNPs are represented by their inherent instability in a biological medium, strategies to both achieve long-term stability and monitor hazardous MNP degradation are needed. We combined a dynamic approach relying on flow field flow fractionation (FFF)-multidetection with conventional techniques to explore frame-by-frame changes of MNPs injected in simulated biological medium, hypothesize the interaction mechanism they are subject to when surrounded by a saline, protein-rich environment, and understand their behaviour at the most critical point of intravenous administration.

The Challenges of O Detection in Biological Fluids: Classical Methods and Translation to Clinical Applications.

Dissolved oxygen (DO) is deeply involved in preserving the life of cellular tissues and human beings due to its key role in cellular metabolism: its alterations may reflect important pathophysiological conditions. DO levels are measured to identify pathological conditions, explain pathophysiological mechanisms, and monitor the efficacy of therapeutic approaches. This is particularly relevant when the measurements are performed in vivo but also in contexts where a variety of biological and synthetic media are used, such as ex vivo organ perfusion.

A Green Analytical Method Combined with Chemometrics for Traceability of Tomato Sauce Based on Colloidal and Volatile Fingerprinting.

Tomato sauce is a world famous food product. Despite standards regulating the production of tomato derivatives, the market suffers frpm fraud such as product adulteration, origin mislabelling and counterfeiting. Methods suitable to discriminate the geographical origin of food samples and identify counterfeits are required.

Application of Af4-Multidetection to Liraglutide in Its Formulation: Preserving and Representing Native Aggregation.

Aggregation is among the most critical parameters affecting the pharmacological and safety profile of peptide Active Pharmaceutical Ingredients (APIs). For this reason, it is of utmost importance to define the exact aggregation state of peptide drugs, particularly when the API is marketed as a ready-to-use solution. Consequently, appropriate non-destructive techniques able to replicate the peptide environment must be employed.

Optimization of a Monobromobimane (MBB) Derivatization and RP-HPLC-FLD Detection Method for Sulfur Species Measurement in Human Serum after Sulfur Inhalation Treatment.

(1) Background: Hydrogen sulfide (HS) is a widely recognized gasotransmitter, with key roles in physiological and pathological processes. The accurate quantification of HS and reactive sulfur species (RSS) may hold important implications for the diagnosis and prognosis of diseases. However, HS species quantification in biological matrices is still a challenge.

Quality Control Platform for the Standardization of a Regenerative Medicine Product.

Adipose tissue is an attractive source of stem cells due to its wide availability. They contribute to the stromal vascular fraction (SVF), which is composed of pre-adipocytes, tissue-progenitors, and pericytes, among others. Because its direct use in medical applications is increasing worldwide, new quality control systems are required.

Synthesis Monitoring, Characterization and Cleanup of Ag-Polydopamine Nanoparticles Used as Antibacterial Agents with Field-Flow Fractionation.

Advances in nanotechnology have opened up new horizons in nanomedicine through the synthesis of new composite nanomaterials able to tackle the growing drug resistance in bacterial strains. Among these, nanosilver antimicrobials sow promise for use in the treatment of bacterial infections. The use of polydopamine (PDA) as a biocompatible carrier for nanosilver is appealing; however, the synthesis and functionalization steps used to obtain Ag-PDA nanoparticles (NPs) are complex and require time-consuming cleanup processes.

Effective Label-Free Sorting of Multipotent Mesenchymal Stem Cells from Clinical Bone Marrow Samples.

Mesenchymal stem cells (MSC) make up less than 1% of the bone marrow (BM). Several methods are used for their isolation such as gradient separation or centrifugation, but these methodologies are not direct and, thus, plastic adherence outgrowth or magnetic/fluorescent-activated sorting is required. To overcome this limitation, we investigated the use of a new separative technology to isolate MSCs from BM; it label-free separates cells based solely on their physical characteristics, preserving their native physical properties, and allows real-time visualization of cells.

FFF-based high-throughput sequence shortlisting to support the development of aptamer-based analytical strategies.

Aptamers are biomimetic receptors that are increasingly exploited for the development of optical and electrochemical aptasensors. They are selected in vitro by the SELEX (Systematic Evolution of Ligands by Exponential Enrichment) procedure, but although they are promising recognition elements, for their reliable applicability for analytical purposes, one cannot ignore sample components that cause matrix effects. This particularly applies when different SELEX-selected aptamers and related truncated sequences are available for a certain target, and the choice of the aptamer should be driven by the specific downstream application.

RNA-seq in DMD urinary stem cells recognized muscle-related transcription signatures and addressed the identification of atypical mutations by whole-genome sequencing.

Urinary stem cells (USCs) are a non-invasive, simple, and affordable cell source to study human diseases. Here we show that USCs are a versatile tool for studying Duchenne muscular dystrophy (DMD), since they are able to address RNA signatures and atypical mutation identification. Gene expression profiling of DMD individuals' USCs revealed a profound deregulation of inflammation, muscle development, and metabolic pathways that mirrors the known transcriptional landscape of DMD muscle and worsens following USCs' myogenic transformation.

A New Predictive Technology for Perinatal Stem Cell Isolation Suited for Cell Therapy Approaches.

The use of stem cells for regenerative applications and immunomodulatory effect is increasing. Amniotic epithelial cells (AECs) possess embryonic-like proliferation ability and multipotent differentiation potential. Despite the simple isolation procedure, inter-individual variability and different isolation steps can cause differences in isolation yield and cell proliferation ability, compromising reproducibility observations among centers and further applications.

An ultracentrifugation - hollow-fiber flow field-flow fractionation orthogonal approach for the purification and mapping of extracellular vesicle subtypes.

In the course of their life span, cells release a multitude of different vesicles in the extracellular matrix (EVs), constitutively and/or upon stimulation, carrying signals either inside or on their membrane for intercellular communication. As a natural delivery tool, EVs present many desirable advantages, such as biocompatibility and low toxicity. However, due to the complex biogenesis of EVs and their high heterogeneity in size distribution and composition, the characterization and quantification of EVs and their subpopulations still represents an enticing analytical challenge.

Unravelling Heterogeneity of Amplified Human Amniotic Fluid Stem Cells Sub-Populations.

Human amniotic fluid stem cells (hAFSCs) are broadly multipotent immature progenitor cells with high self-renewal and no tumorigenic properties. These cells, even amplified, present very variable morphology, density, intracellular composition and stemness potential, and this heterogeneity can hinder their characterization and potential use in regenerative medicine. Celector (Stem Sel ltd.

Perspectives on protein biopolymers: miniaturized flow field-flow fractionation-assisted characterization of a single-cysteine mutated phaseolin expressed in transplastomic tobacco plants.

The development of plant-based protein polymers to employ in biofilm production represents the promising intersection between material science and sustainability, and allows to obtain biodegradable materials that also possess excellent physicochemical properties. A possible candidate for protein biopolymer production is phaseolin, a storage protein highly abundant in P Vulgaris beans. We previously showed that transformed tobacco chloroplasts could be employed to express a mutated phaseolin carrying a signal peptide (directing it into the thylakoids) also enriched of a cysteine residue added to its C-terminal region.

Comprehensive characterization of gold nanoparticles and their protein conjugates used as a label by hollow fiber flow field flow fractionation with photodiode array and fluorescence detectors and multiangle light scattering.

Most of lateral flow immunoassay (LFIA) devices rely on gold nanoparticles (GNP) labeled antibodies or other biospecific proteins, to achieve reagent-less color-based detection. GNP size, GNP-protein conjugation level and its stability are crucial points for the development of precise and accurate methods. In addition, the purification of the GNP-protein conjugates from unreacted protein and GNP, is necessary for adequate analytical performance of the assay.

A new approach for the separation, characterization and testing of potential prionoid protein aggregates through hollow-fiber flow field-flow fractionation and multi-angle light scattering.

Protein misfolding and aggregation are the common mechanisms in a variety of aggregation-dependent diseases. The compromised proteins often assemble into toxic, accumulating amyloid-like structures of various lengths and their toxicity can also be transferred both in vivo and in vitro a prion-like behavior. The characterization of protein interactions, degradation and conformational dynamics in biological systems still represents an analytical challenge in the prion-like protein comprehension.

Widening the Therapeutic Perspectives of Clofazimine by Its Loading in Sulfobutylether β-Cyclodextrin Nanocarriers: Nanomolar IC Values against MDR S. epidermidis.

Clofazimine (CLZ) is an antibiotic with a promising behavior against Gram-positive bacteria; however, the drug is completely insoluble in water and accumulates in fat tissues. We explored nanocarriers, labeled and not labeled with rhodamine, consisting of negatively charged sulfobutylether-β-cyclodextrins for CLZ loading. A new oligomeric carrier was obtained cross-linking βCyD with epichlorohydrin followed by sulfonation in a strongly alkaline aqueous medium.