Publications

Mesenchymal stem cells (MSCs) interact with their surroundings via integrins, which link to the actin cytoskeleton and translate physical cues into biochemical signals through mechanotransduction. N-cadherins enable cell-cell communication and are also linked to the cytoskeleton. This crosstalk between integrins and cadherins modulates MSC mechanotransduction and fate. Here we show the role of this crosstalk in the mechanosensing of viscosity using supported lipid bilayers as substrates of varying viscosity. We functionalize these lipid bilayers with adhesion peptides for integrins (RGD) and N-cadherins (HAVDI), to demonstrate that integrins and cadherins compete for the actin cytoskeleton, leading to an altered MSC mechanosensing response. This response is characterised by a weaker integrin adhesion to the environment when cadherin ligation occurs. We model this competition via a modified molecular clutch model, which drives the integrin/cadherin crosstalk in response to surface viscosity, ultimately controlling MSC lineage commitment. The crosstalk between cell-cell and cell-matrix adhesions regulates stem cell fate. Here, the authors reveal a critical role for matrix viscosity in controlling this crosstalk, which they explain via a modified molecular clutch model.

JTD Keywords: Actin cytoskeleton, Adhesion, Animals, Arginyl-glycyl-aspartic acid, Cadherins, Cell adhesion, Cell communication, Fibronectin, Force transmission, Humans, Hydrogel, Integrins, Lipid bilayers, Matrix, Mechanotransduction, Mechanotransduction, cellular, Mesenchymal stem cells, Mobility, Oligopeptides, Osteogenic differentiation, Substrate stiffness, Vinculin, Viscosity

Degradable polymeric micelles are promising drug delivery systems due to their hydrophobic core and responsive design. When applying micellar nanocarriers for tumor delivery, one of the bottlenecks encountered in vivo is the tumor tissue barrier: crossing the dense mesh of cells and the extracellular matrix (ECM). Sometimes overlooked, the extracellular matrix can trap nanoformulations based on charge, size, and hydrophobicity. Here, we used a simple design of a microfluidic chip with two types of ECM and MCF7 spheroids to allow high-throughput screening of the interactions between biological interfaces and polymeric micelles. To demonstrate the applicability of the chip, a small library of fluorescently labeled polymeric micelles varying in their hydrophilic shell and hydrophobic core forming blocks was studied. Three widely used hydrophilic shells were tested and compared, namely, poly(ethylene glycol), poly(2-ethyl-2-oxazoline), and poly(acrylic acid), along with two enzymatically degradable dendritic hydrophobic cores (based on hexyl or nonyl end groups). Using ratiometric imaging of unimer:micelle fluorescence and FRAP inside the chip model, we obtained the local assembly state and dynamics inside the chip. Notably, we observed different micelle behaviors in the basal lamina ECM, from avoidance of the ECM structure to binding of the poly(acrylic acid) formulations. Binding to the basal lamina correlated with higher uptake into MCF7 spheroids. Overall, we proposed a simple microfluidic chip containing dual ECM and spheroids for the assessment of the interactions of polymeric nanocarriers with biological interfaces and evaluating nanoformulations' capacity to cross the tumor tissue barrier.

JTD Keywords: Extracellular matrix, Microfluidics, Nanoparticle mobility, Polymeric micelles, Tumor-on-a-chip

In order to exploit the pharmacological potential of natural bioactive molecules with low water solubility, such as curcumin, it is necessary to develop formulations, such as amorphous polymer dispersions, which allow a constant release rate and at the same time avoid possible toxicity effects of the crystalline form of the molecule under scrutiny. In this study, polymer dispersions of curcumin were obtained in PADAS, a biodegradable semicrystalline copolymer based on 1,12-dodecanediol, sebacic acid and alanine. The dispersions were fully characterized by means of differential scanning calorimetry and broadband dielectric spectroscopy, and the drug release profile was measured in a simulated body fluid. Amorphous homogeneous binary dispersions were obtained for curcumin mass fraction between 30 and 50%. Curcumin has significantly higher glass transition temperature Tg (≈ 347 K) than the polymer matrix (≈274-277 K depending on the molecular weight), and dispersions displayed Tg's intermediate between those of the pure amorphous components, implying that curcumin acts as an effective antiplasticizer for PADAS. Dielectric spectroscopy was employed to assess the relaxation dynamics of the binary dispersion with 30 wt% curcumin, as well as that of each (amorphous) component separately. The binary dispersion was characterized by a single structural relaxation, a single Johari-Goldstein process, and two local intramolecular processes, one for each component. Interestingly, the latter processes scaled with the Tg of the sample, indicating that they are viscosity-sensitive. In addition, both the pristine polymer and the dispersion exhibited an interfacial Maxwell-Wagner relaxation, likely due to spatial heterogeneities associated with phase disproportionation in this polymer. The release of curcumin from the dispersion in a simulated body fluid followed a Fickian diffusion profile, and 51% of the initial curcumin content was released in 48 h.Copyright © 2023. Published by Elsevier B.V.

JTD Keywords: antioxidant, bioavailability, dielectric spectroscopy, domain havriliak-negami, glass transition temperature, kinetic stability, molecular mobility, nm pores, phase-behavior, physical stability, release kinetics, temperature, thermodynamic quantities, time, Amorphous formulations, Dielectric spectroscopy, Glass transition temperature, Kinetic stability, Kohlrausch-williams-watts, Molecular mobility, Release kinetics

Amorphous molecule-macromolecule mixtures are ubiquitous in polymer technology and are one of the most studied routes for the development of amorphous drug formulations. For these applications it is crucial to understand how the preparation method affects the properties of the mixtures. Here, we employ differential scanning calorimetry and broadband dielectric spectroscopy to investigate dispersions of a small-molecule drug (the Nordazepam anxiolytic) in biodegradable polylactide, both in the form of solvent-cast films and electrospun microfibres. We show that the dispersion of the same small-molecule compound can have opposite (plasticizing or antiplasticizing) effects on the segmental mobility of a biopolymer depending on preparation method, temperature, and polymer enantiomerism. We compare two different chiral forms of the polymer, namely, the enantiomeric pure, semicrystalline L-polymer (PLLA), and a random, fully amorphous copolymer containing both L and D monomers (PDLLA), both of which have lower glass transition temperature (Tg) than the drug. While the drug has a weak antiplasticizing effect on the films, consistent with its higher Tg, we find that it actually acts as a plasticizer for the PLLA microfibres, reducing their Tg by as much as 14 K at 30%-weight drug loading, namely, to a value that is lower than the Tg of fully amorphous films. The structural relaxation time of the samples similarly depends on chemical composition and morphology. Most mixtures displayed a single structural relaxation, as expected for homogeneous samples. In the PLLA microfibres, the presence of crystalline domains increases the structural relaxation time of the amorphous fraction, while the presence of the drug lowers the structural relaxation time of the (partially stretched) chains in the microfibres, increasing chain mobility well above that of the fully amorphous polymer matrix. Even fully amorphous homogeneous mixtures exhibit two distinct Johari–Goldstein relaxation processes, one for each chemical component. Our findings have important implications for the interpretation of the Johari–Goldstein process as well as for the physical stability and mechanical properties of microfibres with small-molecule additives.

JTD Keywords: amorphous pharmaceuticals, beta-relaxation, constant loss, crystallization, dielectric spectroscopy, dynamics, formulation morphology, glass transition, molecular mobility, nanofibers, polylactide, polymer enantiomerism, secondary relaxations, valium metabolite, viscous-liquids, Amorphous pharmaceuticals, Glass-transition, Secondary relaxations

Despite tremendous progress in the understanding of COVID-19, mechanistic insight into immunological, disease-driving factors remains limited. We generated maVie16, a mouse-adapted SARS-CoV-2, by serial passaging of a human isolate. In silico modeling revealed how only three Spike mutations of maVie16 enhanced interaction with murine ACE2. maVie16 induced profound pathology in BALB/c and C57BL/6 mice, and the resulting mouse COVID-19 (mCOVID-19) replicated critical aspects of human disease, including early lymphopenia, pulmonary immune cell infiltration, pneumonia, and specific adaptive immunity. Inhibition of the proinflammatory cyto-kines IFN? and TNF substantially reduced immunopathology. Importantly, genetic ACE2-deficiency completely prevented mCOVID-19 development. Finally, inhalation therapy with recombinant ACE2 fully protected mice from mCOVID-19, revealing a novel and efficient treatment. Thus, we here present maVie16 as a new tool to model COVID-19 for the discovery of new therapies and show that disease severity is determined by cytokine-driven immunopathology and critically dependent on ACE2 in vivo. © Gawish et al.

JTD Keywords: covid-19 mouse model, covid-19 therapy, cytokine storm, immunology, inflammation, mavie16, mouse, mouse-adapted sars-cov-2, program, recombinant soluble ace2, tmprss2, Adaptive immunity, Angiotensin converting enzyme 2, Angiotensin-converting enzyme 2, Animal, Animal cell, Animal experiment, Animal model, Animal tissue, Animals, Apoptosis, Article, Bagg albino mouse, Breathing rate, Bronchoalveolar lavage fluid, C57bl mouse, Cell composition, Cell infiltration, Controlled study, Coronavirus disease 2019, Coronavirus spike glycoprotein, Covid-19, Cytokeratin 18, Cytokine production, Dipeptidyl carboxypeptidase, Disease model, Disease models, animal, Disease severity, Drosophila-melanogaster, Enzyme linked immunosorbent assay, Expression vector, Flow cytometry, Gamma interferon, Gene editing, Gene expression, Gene mutation, Genetic engineering, Genetics, Glycosylation, High mobility group b1 protein, Histology, Histopathology, Immune response, Immunocompetent cell, Immunology, Immunopathology, Interferon-gamma, Interleukin 2, Metabolism, Mice, inbred balb c, Mice, inbred c57bl, Mouse-adapted sars-cov-2, Myeloperoxidase, Neuropilin 1, Nonhuman, Nucleocapsid protein, Pathogenicity, Peptidyl-dipeptidase a, Pyroptosis, Recombinant soluble ace2, Renin angiotensin aldosterone system, Rna extraction, Rna isolation, Sars-cov-2, Severe acute respiratory syndrome coronavirus 2, Spike glycoprotein, coronavirus, T lymphocyte activation, Trabecular meshwork, Tumor necrosis factor, Virology, Virus load, Virus replication, Virus transmission, Virus virulence

Most lectins bind carbohydrate ligands with relatively low affinity, making the identification of optimal ligands challenging. Here we introduce a point accumulation in nanoscale topography (PAINT) super-resolution microscopy method to capture weak glycan-lectin interactions at the single-molecule level in living cells (Glyco-PAINT). Glyco-PAINT exploits weak and reversible sugar binding to directly achieve single-molecule detection and quantification in cells and is used to establish the relative kon and koff rates of a synthesized library of carbohydrate-based probes, as well as the diffusion coefficient of the receptor-sugar complex. Uptake of ligands correlates with their binding affinity and residence time to establish structure-function relations for various synthetic glycans. We reveal how sugar multivalency and presentation geometry can be optimized for binding and internalization. Overall, Glyco-PAINT represents a powerful approach to study weak glycan-lectin interactions on the surface of living cells, one that can be potentially extended to a variety of lectin-sugar interactions.© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.

JTD Keywords: dc-sign, density, dimerization, endocytosis, lateral mobility, ligand-binding, mannose receptor, proteins, recognition, Animal, Animals, Cell membrane, Cell membrane permeability, Chemistry, Cho cell line, Cho cells, Cricetulus, Cysteine-rich domain, Kinetics, Lectin, Lectins, Ligand, Ligands, Molecular library, Multivariate analysis, Polysaccharide, Polysaccharides, Procedures, Protein binding, Single molecule imaging, Small molecule libraries, Structure activity relation, Structure-activity relationship

We characterize amorphous solid dispersions (ASDs) of the Chloramphenicol antibiotic in two biodegradable polylactic acid polymers, namely a commercial sample of enantiomeric pure PLLA and a home-synthesized PDLLA copolymer, investigating in particular the effect of polylactic acid in stabilizing the amorphous form of the drug and controlling its release (e.g. for antitumoral purposes). Broadband dielectric spectroscopy and differential scanning calorimetry are employed to study the homogeneity, glass transition temperature and relaxation dynamics of solvent-casted ASD membranes with different drug concentrations. We observe improved physical stability of the ASDs with respect to the pure drug, as well as a plasticizing effect of the antibiotic on the polymer, well described by the Gordon-Taylor equation. The release of the active pharmaceutical ingredient from the films in a simulated body fluid is studied by UV/vis spectroscopy at two different drug concentrations (5 and 20% in weight). The amount of released drug is found to be proportional to the square root of time, with proportionality constant that is almost the same in both dispersions, despite the fact that the relaxation time and thus the viscosity of the two samples differ by four orders of magnitude at body temperature. Since the drug release kinetics does not display a significant dependence on the drug content in the carrier, it may be expected to remain roughly constant during longer release times.

JTD Keywords: Amorphous drug, Controlled liberation, Dielectric spectroscopy, Molecular mobility, Plasticizer, Polymer enantiomerism

Extracellular matrix (ECM) proteins are key mediators of cell/material interactions. The surface density and conformation of these proteins adsorbed on the material surface influence cell adhesion and the cellular response. We have previously shown that subtle variations in surface chemistry lead to drastic changes in the conformation of adsorbed fibronectin (FN). On poly(ethyl acrylate) (PEA), FN unfolds and displays domains for cell adhesion and FN-FN interaction, whereas on poly(methyl acrylate) (PMA) – with only one methyl group less – FN remains globular as it is in solution. The effect of the strength of the protein/material interaction in cell response, and its relation to protein density and conformation, has received limited attention so far. In this work, we used FN-functionalized AFM cantilevers to evaluate, via force spectroscopy, the strength of interaction between fibronectin and the underlying polymer which controls FN conformation (PEA and PMA). We found that the strength of FN/PEA interaction is significantly higher than FN/PMA, which limits the mobility of FN layer on PEA, reduces the ability of cells to mechanically reorganize FN and then leads to enhanced proteolysis and degradation of the surrounding matrix with compromised cell viability. By contrast, both PEA and PMA support cell adhesion when FN density is increased and also in the presence of serum or other serum proteins, including vitronectin (VN) and bovine serum albumin (BSA), which provide a higher degree of mobility to the matrix. Statement of Significance: The identification of parameters influencing cell response is of paramount importance for the design of biomaterials that will act as synthetic scaffolds for cells to anchor, grow and, eventually, become specialised tissues. Cells interact with materials through an intermediate layer of proteins adsorbed on the material surface. It is known that the density and conformation of these proteins determine cell behaviour. Here we show that the strength of protein/material interactions, which has received very limited attention so far, is key to understand the cellular response to biomaterials. Very strong protein/material interactions reduce the ability of cells to mechanically reorganize proteins at the material interface which results in enhanced matrix degradation, leading ultimately to compromised cell viability.

JTD Keywords: Fibronectin adsorption, Fibronectin remodeling, Protein mobility, Protein-material interaction strength

Existing analytical techniques used for the determination of cannabinoids in Cannabis sativa L. (Cannabis) plants mostly rely on chromatography-based methods. As a rapid alternative for the direct analysis of them, thermal desorption (TD)-ion mobility spectrometry (IMS) was used for obtaining spectral fingerprints of single cannabinoids from Cannabis plant extracts and from plant residues on hands after their manipulation. The ionization source was 63Ni, with automatic switchable polarity. Although in both ionization modes there were signals in the TD-IMS spectra of the plant extracts and residues that could be assigned to concrete cannabinoids and chemotypes, most of them could not be clearly distinguished. Alternatively, the global spectral data of the plant extracts and residues were pre-processed and then, using principal component analysis (PCA)-linear discriminant analysis (LDA), grouped in function of their chemotype in a more feasible way. Using this approach, the possibility of false positive responses was also studied analyzing other non-Cannabis plants and tobacco, which were clustered in a different group to those of Cannabis. Therefore, TD-IMS, as analytical tool, and PCA-LDA, as a strategy for data reduction and pattern recognition, can be applied for on-site chemotaxonomic discrimination of Cannabis varieties and detection of illegal marijuana since the IMS equipment is portable and the analysis time is highly short.

JTD Keywords: Cannabis sativa L., Cannabinoids, Chemometrics, ChemotypeIon mobility spectrometry

Abstract Blind Source Separation (BSS) techniques aim to extract a set of source signals from a measured mixture in an unsupervised manner. In the chemical instrumentation domain source signals typically refer to time-varying analyte concentrations, while the measured mixture is the set of observed spectra. Several techniques exist to perform BSS on Ion Mobility Spectrometry, being Simple-to-use interactive self-modeling mixture analysis (SIMPLISMA) and Multivariate Curve Resolution (MCR) the most commonly used. The addition of a multi-capillary gas chromatography column using the ion mobility spectrometer as detector has been proposed in the past to increase chemical resolution. Short chromatography times lead to high levels of co-elution, and ion mobility spectra are key to resolve them. For the first time, BSS techniques are used to deconvolve samples of the gas chromatography - ion mobility spectrometry tandem. We propose a method to extract spectra and concentration profiles based on the application of MCR in a sliding window. Our results provide clear concentration profiles and pure spectra, resolving peaks that were not detected by the conventional use of MCR. The proposed technique could also be applied to other hyphenated instruments with similar strong co-elutions.

JTD Keywords: Blind Source Separation, Multivariate Curve Resolution, Ion Mobility Spectrometry, Gas Chromatography, Hyphenated instrumentation, SIMPLISMA, co-elution

The performance of three different types of ion mobility spectrometer (IMS) devices: GDA2 with a radioactive ion source (Airsense, Germany), UV-IMS with a photo-ionization source (G.A.S. Germany) and VG-Test with a corona discharge source (3QBD, Israel) was studied. The gas-phase ion chemistry in the IMS devices affected the species formed and their measured reduced mobility values. The sensitivity and limit of detection for trimethylamine (TMA), putrescine and cadaverine were compared by continuous monitoring of a stream of air with a given concentration of the analyte and by measurement of headspace vapors of TMA in a sealed vial. Preprocessing of the mobility spectra and the effectiveness of multivariate curve resolution techniques (MCR-LASSO) improved the accuracy of the measurements by correcting baseline effects and adjusting for variations in drift time as well as enhancing the signal to noise ratio and deconvolution of the complex data matrix to their pure components. The limit of detection for measurement of the biogenic amines by the three IMS devices was between 0.1 and 1.2 ppm (for TMA with the VG-Test and GDA, respectively) and between 0.2 and 0.7 ppm for putrescine and cadaverine with all three devices. Considering the uncertainty in the LOD determination there is almost no statistically significant difference between the three devices although they differ in their operating temperature, ionization method, drift tube design and dopant chemistry. This finding may have general implications on the achievable performance of classic IMS devices.

JTD Keywords: Biogenic amines, Comparison of performance, Ion mobility spectrometry, Sensitivity, Signal processing, Vapor concentration

The off-flavor of “tainted wine” is attributed mainly to the presence of 2,4,6-trichloroanisole (2,4,6-TCA) in the wine. In the present study the atmospheric pressure gas-phase ion chemistry, pertaining to ion mobility spectrometry, of 2,4,6-trichloroanisole was investigated. In positive ion mode the dominant species is a monomer ion with a lower intensity dimer species with reduced mobility values (K0) of 1.58 and 1.20 cm2 V−1 s−1, respectively. In negative mode the ion with K0 = 1.64 cm2 V−1 s−1 is ascribed to a trichlorophenoxide species while the ions with K0 = 1.48 and 1.13 cm2 V−1 s−1 are attributed to chloride attachment adducts of a TCA monomer and dimer, respectively. The limit of detection of the system for 2,4,6-TCA dissolved in dichloromethane deposited on a filter paper was 2.1 ug and 1.7 ppm in the gas phase. In ethanol and in wine the limit of detection is higher implying that pre-concentration and pre-separation are required before IMS can be used to monitor the level of TCA in wine.

JTD Keywords: 2,4,6-Trichloroanisole, Gas phase ion chemistry, Ion mobility spectrometry, "Tainted wine"

Background: Sepsis is one of the main causes of death in adult intensive care units. The major drawbacks of the different methods used for its diagnosis and monitoring are their inability to provide fast responses and unsuitability for bedside use. In this study, performed using a rat sepsis model, we evaluate breath analysis with Ion Mobility Spectrometry (IMS) as a fast, portable and non-invasive strategy. Methods: This study was carried out on 20 Sprague-Dawley rats. Ten rats were injected with lipopolysaccharide from Escherichia coli and ten rats were IP injected with regular saline. After a 24-h period, the rats were anaesthetized and their exhaled breaths were collected and measured with IMS and SPME-gas chromatography/mass spectrometry (SPME-GC/MS) and the data were analyzed with multivariate data processing techniques. Results: The SPME-GC/MS dataset processing showed 92% accuracy in the discrimination between the two groups, with a confidence interval of between 90.9% and 92.9%. Percentages for sensitivity and specificity were 98% (97.5–98.5%) and 85% (84.6–87.6%), respectively. The IMS database processing generated an accuracy of 99.8% (99.7–99.9%), a specificity of 99.6% (99.5–99.7%) and a sensitivity of 99.9% (99.8–100%). Conclusions: IMS involving fast analysis times, minimum sample handling and portable instrumentation can be an alternative for continuous bedside monitoring. IMS spectra require data processing with proper statistical models for the technique to be used as an alternative to other methods. These animal model results suggest that exhaled breath can be used as a point-of-care tool for the diagnosis and monitoring of sepsis.

JTD Keywords: Sepsis, Volatile organic compounds, Ion mobility spectrometer, Rat model, Bedside patient systems, Non-invasive detection

In this work, a new methodology to analyze spectra time-series obtained from ion mobility spectrometry (IMS) has been investigated. The proposed method combines the advantages of multivariate curve resolution-alternating least squares (MCR-ALS) for an optimal physical and chemical interpretation of the system (qualitative information) and a multivariate calibration technique such as polynomial partial least squares (poly-PLS) for an improved quantification (quantitative information) of new samples. Ten different concentrations of 2-butanone and ethanol were generated using a volatile generator based on permeation tubes. The different concentrations were measured with IMS. These data present a non-linear behaviour as substance concentration increases. Although MCR-ALS is based on a bilinear decomposition, non-linear behaviour can be modelled adding new components to the model. After spectral pre-processing, MCR-ALS was applied aiming to get information about the ionic species that appear in the drift tube and their evolution with the analyte concentration. By resolving the IMS data matrix, concentration profiles and pure spectra of the different ionic species have been obtained for both analytes. Finally, poly-PLS was used in order to build a calibration model using concentration profiles obtained from MCR-ALS for ethanol and 2-butanone. The results, with more than 99% of explained variance for both substances, show the feasibility of using MCR-ALS to resolve IMS datasets. Furthermore, similar or better prediction accuracy is achieved when concentration profiles from MCR-ALS are used to build a calibration model (using poly-PLS) compared to other standard univariate and multivariate calibration methodologies.

JTD Keywords: Ion Mobility Spectrometry, Multivariate Curve Resolution, Gas phase ion chemistry, Multivariate calibration

The potential of a vanguard technique as is the Ion Mobility Spectrometry with Ultraviolet ionization (UV-IMS) coupled to a Continuous Flow System (CFS) have been demonstrated in this work by using a Gas Phase Separator (GPS). This vanguard system (CFS-GPS-UV-IMS) has been used for the analysis of different types of white wines to obtain a characteristic profile for each type of wine and their posterior classification using different chemometric tools. Precision of the method was 3.1% expressed as relative standard deviation. A deep chemometric study was carried out for the classification of the four types of wines selected. The best classification performance was obtained by first reducing the data dimensionality by Principal Component Analysis (PCA) followed by Linear Discriminant Analysis (LDA) and finally using a K-Nearest Neighbour (kNN) classifier. The classification rate in an independent validation set were 92.0% classification rate value with confidence interval [89.0%, 95.0%] at P = 0.05 confidence level. The same white wines analyzed by using CFS-GPS-UV-IMS were analyzed by using Gas Chromatography with a Flame Detector (GC-FID) as conventional technique. The chromatographic method used for the determination of superior alcohols in wine samples shown in the Regulation CEE 1238/1992 was selected to carry out the analysis of the same samples set and later the classification using appropriate chemometric tools. In this case, strategies PCA-LDA and kNN classifier were also used for the correct classification of the wine samples. This combination showed similar results to the ones obtained with the proposed method.

JTD Keywords: Classification, White wines, Ultraviolet-Ion Mobility Spectrometry, Gas Phase Separate, Vanguard method, Continuous Flow System, Chemometric analysis.

Multivariate Curve Resolution (MCR) aims to blindly recover the concentration profile and the source spectra without any prior supervised calibration step. It is well known that imposing additional constraints like positiveness, closure and others may improve the quality of the solution. When a physico-chemical model of the process is known, this can be also introduced constraining even more the solution. In this paper, we apply MCR to Ion Mobility Spectra. Since instrumental models suggest that peaks are of Gaussian shape with a width depending on the instrument resolution, we introduce that each source is characterized by a linear superposition of Gaussian peaks of fixed spread. We also prove that this model is able to fit wider peaks departing from pure Gaussian shape. Instead of introducing a non-linear Gaussian peak fitting, we use a very dense model and rely on a least square solver with L1-norm regularization to obtain a sparse solution. This is accomplished via Least Absolute Shrinkage and Selection Operator (LASSO). Results provide nicely resolved concentration profiles and spectra improving the results of the basic MCR solution.

JTD Keywords: Blind source separation, Ion mobility spectrometry, Multivariate curve resolution, Sparse solution, Non negative matrix factorization

Proteins of the Hha/YmoA family co-regulate with H-NS the expression of virulence factors in Enterobacteriaceae. Vibrio cholerae lacks Hha-like proteins and its H-NS (vcH-NS) is unable to bind Hha, in spite of the conservation of a key residue for Hha binding by Escherichia coli H-NS (ecH-NS). Exchange of the residues in position 9 between vcH-NS and ecH-NS strongly reduces Hha binding by ecH-NS and introduces it in vcH- NS. These mutations strongly affect the repression of the hemolysin operon in E. coli and the electrophoretic mobility of complexes formed with a DNA fragment containing its regulatory region.

JTD Keywords: Nucleoid associated protein, H-NS, Hha, Transcription repression, NMR, Electrophoretic mobility shift assays

Miniaturization is a powerful trend for smart chemical instrumentation in a diversity of applications.. It is know that miniaturization in IMS leads to a degradation of the system characteristics. For the present work, we are interested in signal processing solutions to mitigate limitations introduced by limited drift tube length that basically involve a loss of chemical selectivity. While blind source separation techniques (BSS) are popular in other domains, their application for smart chemical instrumentation is limited. However, in some conditions, basically linearity, BSS may fully recover the concentration time evolution and the pure spectra with few underlying hypothesis. This is extremely helpful in conditions where non-expected chemical interferents may appear, or unwanted perturbations may pollute the spectra. SIMPLISMA has been advocated by Harrington et al. in several papers. However, more modem methods of BSS for bilinear decomposition with the restriction of positiveness have appeared in the last decade. In order to explore and compare the performances of those methods a series of experiments were performed.

JTD Keywords: Ion Mobility Spectrometry (IMS), Blind Source Separation (BSS), Multivariate Analysis, SIMPLISMA, MCR, Non-Negative Matrix Factorization (NMF)

This work presents the possibilities offered by a blind source separation method such Multivariate Curve Resolution- Alternating Least Squares (MCR-ALS) in the analysis of Ion Mobility Spectra (IMS). Two security applications are analyzed in this context: the detection of TNT both in synthetic and real samples. Results obtained show the possibilities offered by the direct analysis of the drift time spectra when an appropriate resolution method is used.

JTD Keywords: Ion Mobility Spectrometry, Multivariate Curve Resolution, Security, LIMS, MCR-ALS