Quantification of Silymarin in Silybi mariani fructus: Challenging the Analytical Performance of Benchtop vs. Handheld NIR Spectrometers on Whole Seeds.

The content of the flavonolignan mixture silymarin and its individual components (silichristin, silidianin, silibinin A, silibinin B, isosilibinin A, and isosilibinin B) in whole and milled milk thistle seeds (Silybi mariani fructus) was analyzed with near-infrared spectroscopy. The analytical performance of one benchtop and two handheld near-infrared spectrometers was compared. Reference analysis was performed with HPLC following a Soxhlet extraction (European Pharmacopoeia) and a more resource-efficient ultrasonic extraction. The reliability of near-infrared spectral analysis determined through partial least squares regression models constructed independently for the spectral datasets obtained by the three spectrometers was as follows. The benchtop device NIRFlex N-500 performed the best both for milled and whole seeds with a root mean square error of CV between 0.01 and 0.17%. The handheld spectrometer MicroNIR 2200 as well as the microPHAZIR provided a similar performance (root mean square error of CV between 0.01 and 0.18% and between 0.01 and 0.23%, respectively). We carried out quantum chemical simulation of near-infrared spectra of silichristin, silidianin, silibinin, and isosilibinin for interpretation of the results of spectral analysis. This provided understanding of the absorption regions meaningful for the calibration. Further, it helped to better separate how the chemical and physical properties of the samples affect the analysis. While the study demonstrated that milling of samples slightly improves the performance, it was deemed to be critical only for the analysis carried out with the microPHAZIR. This study evidenced that rapid and nondestructive quantification of silymarin and individual flavonolignans is possible with miniaturized near-infrared spectroscopy in whole milk thistle seeds.

Challenging handheld NIR spectrometers with moisture analysis in plant matrices: Performance of PLSR vs. GPR vs. ANN modelling.

The global demand for natural products grows rapidly, intensifying the request for the development of high-throughput, fast, non-invasive tools for quality control applicable on-site. Moisture content is one of the most important quality parameters of natural products. It determines their market suitability, stability and shelf life and should preferably be constantly monitored. Miniaturized near-infrared (NIR) spectroscopy is a powerful method for on-site analysis, potentially fulfilling this requirement. Here, a feasibility study for applicability and analytical performance of three miniaturized NIR spectrometers and two benchtop instruments was evaluated in that scenario. The case study involved 192 dried plant extracts composed of five different plants harvested in different countries at various times within two years. The reference analysis by Karl Fischer titration determined the water content in this sample set between 1.36% and 6.47%. For the spectroscopic analysis half of the samples were laced with a drying agent to comply with the industry standard. The performance of various calibration models for NIR analysis was evaluated on the basis of root-mean square error of prediction (RMSEP) determined for an independent test set. Partial least squares regression (PLSR), Gaussian process regression (GPR) and artificial neural network (ANN) models were constructed for the spectral sets from each instrument. GPR and ANN models performed superior for all samples measured by handheld spectrometers and for native ones analyzed by benchtop instruments. Moreover, the accuracy penalty when analyzing native samples was lower for GPR and ANN prediction as well. With GPR or ANN calibration, miniaturized spectrometers offered the prediction performance at the level of the benchtop instruments. Therefore, in this analytical application miniaturized spectrometers can be used on-site with no penalty to the performance vs. laboratory-based NIR analysis.

BIN2 orchestrates platelet calcium signaling in thrombosis and thrombo-inflammation.

Store-operated Ca2+ entry (SOCE) is the major route of Ca2+ influx in platelets. The Ca2+ sensor stromal interaction molecule 1 (STIM1) triggers SOCE by forming punctate structures with the Ca2+ channel Orai1 and the inositol trisphosphate receptor (IP3R), thereby linking the endo-/sarcoplasmic reticulum to the plasma membrane. Here, we identified the BAR domain superfamily member bridging integrator 2 (BIN2) as an interaction partner of STIM1 and IP3R in platelets. Deletion of platelet BIN2 (Bin2fl/fl,Pf4-Cre mice) resulted in reduced Ca2+ store release and Ca2+ influx in response to all tested platelet agonists. These defects were a consequence of impaired IP3R function in combination with defective STIM1-mediated SOC channel activation, while Ca2+ store content and agonist-induced IP3 production were unaltered. This severely defective Ca2+ signaling translated into impaired thrombus formation under flow and a protection of Bin2fl/fl,Pf4-Cre mice in models of arterial thrombosis and stroke. Our results establish BIN2 as a central regulator of platelet activation in thrombosis and thrombo-inflammatory disease settings.

Principles and Applications of Vibrational Spectroscopic Imaging in Plant Science: A Review.

Detailed knowledge about plant chemical constituents and their distributions from organ level to sub-cellular level is of critical interest to basic and applied sciences. Spectral imaging techniques offer unparalleled advantages in that regard. The core advantage of these technologies is that they acquire spatially distributed semi-quantitative information of high specificity towards chemical constituents of plants. This forms invaluable asset in the studies on plant biochemical and structural features. In certain applications, non-invasive analysis is possible. The information harvested through spectral imaging can be used for exploration of plant biochemistry, physiology, metabolism, classification, and phenotyping among others, with significant gains for basic and applied research. This article aims to present a general perspective about vibrational spectral imaging/micro-spectroscopy in the context of plant research. Within the scope of this review are infrared (IR), near-infrared (NIR) and Raman imaging techniques. To better expose the potential and limitations of these techniques, fluorescence imaging is briefly overviewed as a method relatively less flexible but particularly powerful for the investigation of photosynthesis. Included is a brief introduction to the physical, instrumental, and data-analytical background essential for the applications of imaging techniques. The applications are discussed on the basis of recent literature.

Loss of Orai2-Mediated Capacitative Ca2+ Entry Is Neuroprotective in Acute Ischemic Stroke.

Background and Purpose- Ischemic stroke is one of the leading causes of disability and death. The principal goal of acute stroke treatment is the recanalization of the occluded cerebral arteries, which is, however, only effective in a very narrow time window. Therefore, neuroprotective treatments that can be combined with recanalization strategies are needed. Calcium overload is one of the major triggers of neuronal cell death. We have previously shown that capacitative Ca2+ entry, which is triggered by the depletion of intracellular calcium stores, contributes to ischemia-induced calcium influx in neurons, but the responsible Ca2+ channel is not known. Methods- Here, we have generated mice lacking the calcium channel subunit Orai2 and analyzed them in experimental stroke. Results- Orai2-deficient mice were protected from ischemic neuronal death both during acute ischemia under vessel occlusion and during ischemia/reperfusion upon successful recanalization. Calcium signals induced by calcium store depletion or oxygen/glucose deprivation were significantly diminished in Orai2-deficient neurons demonstrating that Orai2 is a central mediator of neuronal capacitative Ca2+ entry and is involved in calcium overload during ischemia. Conclusions- Our experimental data identify Orai2 as an attractive target for pharmaceutical intervention in acute stroke.

At-Line Monitoring of the Extraction Process of Rosmarini Folium via Wet Chemical Assays, UHPLC Analysis, and Newly Developed Near-Infrared Spectroscopic Analysis Methods.

The present study demonstrates the applicability of at-line monitoring of the extraction process of Rosmarinus officinalis L. leaves (Rosmarini folium) and the development of near-infrared (NIR) spectroscopic analysis methods. Therefore, whole dried Rosmarini folium samples were extracted by maceration with 70% (v/v) ethanol. For the experimental design three different specimen-taking plans were chosen. At first, monitoring was carried out using three common analytical methods: (a) total hydroxycinnamic derivatives according to the European Pharmacopoeia, (b) total phenolic content according to Folin-Ciocalteu, and (c) rosmarinic acid content measured by UHPLC-UV analysis. Precision validation of the wet chemical assays revealed a repeatability of (a) 0.12% relative standard deviation (RSD), (b) 1.1% RSD, and (c) 0.28% RSD, as well as an intermediate precision of (a) 4.1% RSD, (b) 1.3% RSD, and (c) 0.55% RSD. The collected extracts were analyzed with a NIR spectrometer using a temperature-controlled liquid attachment. Samples were measured in transmission mode with an optical path length of 1 mm. The combination of the recorded spectra and the previously obtained analytical reference values in conjunction with multivariate data analysis enabled the successful establishment of partial least squares regression (PLSR) models. Coefficients of determination (R2) were: (a) 0.94, (b) 0.96, and (c) 0.93 (obtained by test-set validation). Since Pearson correlation analysis revealed that the reference analyses correlated with each other just one of the PSLR models is required. Therefore, it is suggested that PLSR model (b) be used for monitoring the extraction process of Rosmarini folium. The application of NIR spectroscopy provides a fast and non-invasive alternative analysis method, which can subsequently be implemented for on- or in-line process control.

Information and order of information effects on consumers' acceptance and valuation for genetically modified edamame soybean.

This study examines two different strategies with respect to managing the order in which information about genetically modified (GM) technology would reach and impact consumers of edamame, often referred to as the "vegetable soybean". Edamame are soybeans harvested while the beans are young and soft. We capture consumers' willingness to pay (WTP) for unlabeled edamame, non-GM edamame, and GM edamame using a non-hypothetical random nth price auction. We elicit consumers' valuation for each edamame product before and after introducing information, and test two strategies where the order of providing positive and negative information is reversed. The results suggest that negative information affects WTP to a much greater extent than positive information. Hence a strategy to proactively deal with eventual negative press about GM technology did not lead to a different result than a strategy that would react to or attempt to thwart negative information with positive information at a later date. These findings suggest that it would be difficult to introduce new GM edamame as edible products in the market as marginally negative preconceptions about GM at the time of the experiment were easier to reinforce with negative information than to combat with positive information about GM.

Near-infrared and Mid-infrared Spectroscopic Techniques for a Fast and Nondestructive Quality Control of Thymi herba.

In this study, novel near-infrared and attenuated total reflectance mid-infrared spectroscopic methods coupled with multivariate data analysis were established enabling the determination of thymol, rosmarinic acid, and the antioxidant capacity of Thymi herba. A new high-performance liquid chromatography method and UV-Vis spectroscopy were applied as reference methods. Partial least squares regressions were carried out as cross and test set validations. To reduce systematic errors, different data pretreatments, such as multiplicative scatter correction, 1st derivative, or 2nd derivative, were applied on the spectra. The performances of the two infrared spectroscopic techniques were evaluated and compared. In general, attenuated total reflectance mid-infrared spectroscopy demonstrated a slightly better predictive power (thymol: coefficient of determination = 0.93, factors = 3, ratio of performance to deviation = 3.94; rosmarinic acid: coefficient of determination = 0.91, factors = 3, ratio of performance to deviation = 3.35, antioxidant capacity: coefficient of determination = 0.87, factors = 2, ratio of performance to deviation = 2.80; test set validation) than near-infrared spectroscopy (thymol: coefficient of determination = 0.90, factors = 6, ratio of performance to deviation = 3.10; rosmarinic acid: coefficient of determination = 0.92, factors = 6, ratio of performance to deviation = 3.61, antioxidant capacity: coefficient of determination = 0.91, factors = 6, ratio of performance to deviation = 3.42; test set validation). The capability of infrared vibrational spectroscopy as a quick and simple analytical tool to replace conventional time and chemical consuming analyses for the quality control of T. herba could be demonstrated.

A Cdc42/RhoA regulatory circuit downstream of glycoprotein Ib guides transendothelial platelet biogenesis.

Blood platelets are produced by large bone marrow (BM) precursor cells, megakaryocytes (MKs), which extend cytoplasmic protrusions (proplatelets) into BM sinusoids. The molecular cues that control MK polarization towards sinusoids and limit transendothelial crossing to proplatelets remain unknown. Here, we show that the small GTPases Cdc42 and RhoA act as a regulatory circuit downstream of the MK-specific mechanoreceptor GPIb to coordinate polarized transendothelial platelet biogenesis. Functional deficiency of either GPIb or Cdc42 impairs transendothelial proplatelet formation. In the absence of RhoA, increased Cdc42 activity and MK hyperpolarization triggers GPIb-dependent transmigration of entire MKs into BM sinusoids. These findings position Cdc42 (go-signal) and RhoA (stop-signal) at the centre of a molecular checkpoint downstream of GPIb that controls transendothelial platelet biogenesis. Our results may open new avenues for the treatment of platelet production disorders and help to explain the thrombocytopenia in patients with Bernard-Soulier syndrome, a bleeding disorder caused by defects in GPIb-IX-V.

Application of benchtop and portable near-infrared spectrometers for predicting the optimum harvest time of Verbena officinalis.

This study examined the applicability of near-infrared (NIR) spectroscopy coupled with multivariate data analysis (MVA) to determine the ideal harvest time of Verbena officinalis. NIR analyses were performed non-invasively on the fresh plant material based on the quantification of the key constituents verbenalin and verbascoside. Vibrational spectroscopic measurements were performed applying a conventional NIR benchtop device as well as a laboratory independent handheld NIR spectrometer. A novel high performance liquid chromatography (HPLC) method was applied as a reference method. For both instruments partial least squares (PLS) regression models were established performing cross validations (CV) and test-set validations (TSV). Quality parameters obtained for the benchtop device revealed that the newly established NIR method enabled reliable quantifications of the main compounds verbenalin and verbascoside related to the dried and fresh plant material. The results of the miniaturised spectrometer revealed that accurate quantitative calibration models could be developed for verbascoside achieving a comparable prediction power to the benchtop device. PLS models for verbenalin were less precise suggesting the application of portable devices including a different spectral range and resolution. The work demonstrated the feasibility of NIR vibrational spectroscopy performing direct measurements on pharmaceutically relevant fresh plant material enabling a quick and simple determination of the ideal harvest time of Verbena officinalis.

FcγRIIB on liver sinusoidal endothelial cells is essential for antibody-induced GPVI ectodomain shedding in mice.

The activating platelet collagen receptor glycoprotein VI (GPVI) is a promising antithrombotic target because of its central role in arterial thrombosis and its minor relevance for normal hemostasis. The receptor can be specifically targeted by antibodies and irreversibly downregulated in circulating platelets in vivo, resulting in long-term antithrombotic protection in mice. This GPVI immunodepletion predominantly occurs through ectodomain shedding, which is accompanied by a transient drop in peripheral platelet counts. Mechanistic studies on this targeted GPVI loss have been hampered because it cannot be reproduced in isolated platelets in vitro. Here we show that both the transient thrombocytopenia and GPVI ectodomain shedding depend on the Fc portion of the anti-GPVI antibody and its interaction with the inhibitory Fcγ receptor (FcγR)IIB. In wild-type, but not Fcgr2b(-/-) mice, anti-GPVI-opsonized platelets became transiently trapped in the liver followed by the appearance of the soluble GPVI ectodomain in the plasma. Depletion of Kupffer cells neither affected anti-GPVI-induced platelet accumulation nor GPVI shedding, demonstrating that the other major FcγRIIB-expressing cell type, liver sinusoidal endothelial cells, is required for both processes to occur. These results reveal a novel and unexpected function of hepatic FcγRIIB in the targeted downregulation of GPVI in vivo.

Normal Platelet Integrin Function in Mice Lacking Hydrogen Peroxide-Induced Clone-5 (Hic-5).

Integrin αIIbß3 plays a central role in the adhesion and aggregation of platelets and thus is essential for hemostasis and thrombosis. Integrin activation requires the transmission of a signal from the small cytoplasmic tails of the α or ß subunit to the large extracellular domains resulting in conformational changes of the extracellular domains to enable ligand binding. Hydrogen peroxide-inducible clone-5 (Hic-5), a member of the paxillin family, serves as a focal adhesion adaptor protein associated with αIIbß3 at its cytoplasmic tails. Previous studies suggested Hic-5 as a novel regulator of integrin αIIbß3 activation and platelet aggregation in mice. To assess this in more detail, we generated Hic-5-null mice and analyzed activation and aggregation of their platelets in vitro and in vivo. Surprisingly, lack of Hic-5 had no detectable effect on platelet integrin activation and function in vitro and in vivo under all tested conditions. These results indicate that Hic-5 is dispensable for integrin αIIbß3 activation and consequently for arterial thrombosis and hemostasis in mice.

Trust and perception related to information about biofuels in Belgium.

Public knowledge regarding green energy is limited and there is a perceived lack of information, particularly in the case of biofuels. This study investigates the information seeking behaviour of fuel consumers with regard to biofuels, their preferences for particular elements and channels of information and their trust in information sources. The majority of respondents were interested in many elements of information, and particularly in the tax (dis)advantages associated with biofuels. They prefer to obtain information via newspapers and brochures. Scientists, environmental and consumer organizations and the government are perceived as the most trustworthy sources of information whilst word of mouth sources, journalists and the fuel sector are less well trusted. Four consumer clusters could be distinguished, on the basis of the perceived relative trustworthiness of information sources, and these clusters have significantly different socio-demographic characteristics and preferences for elements and channels of information.

Determination of carbohydrates in medicinal plants--comparison between TLC, mf-MELDI-MS and GC-MS.

INTRODUCTION: Quality control in the pharmaceutical and phytopharmaceutical industries requires fast and reliable methods for the analysis of raw materials and final products. OBJECTIVE: This study evaluates different analytical approaches in order to recognise the most suitable technique for the analysis of carbohydrates in herbal drug preparations. METHODOLOGY: The specific focus of the study is on thin-layer chromatography (TLC), gas chromatography (GC), and a newly developed mass spectrometric method, i.e. matrix free material enhanced laser desorption/ionisation time of flight mass spectrometry (mf-MELDI-MS). Samples employed in the study were standards and microwave-assisted water extracts from Quercus. RESULTS: TLC analysis proved the presence of mono-, di- and trisaccharides within the biological sample and hinted at the existence of an unknown carbohydrate of higher oligomerisation degree. After evaluation of different derivatisation techniques, GC-MS confirmed data obtained via TLC for mono- to trisaccharides, delivering additionally quantified values under a considerable amount of time. A carbohydrate of higher oligomerisation degree could not be found. The application of mf-MELDI-MS further confirmed the presence of carbohydrates up to trisaccharides, also hinting at the presence of a form of tetrasaccharide. Besides this information, mf-MELDI-MS delivered further data about other substances present in the extract. Quantitative determination resulted in 1.750, 1.736 and 0.336 mg/mL for glucose, sucrose and raffinose respectively. CONCLUSION: Evaluation of all three techniques employed, clearly proved the heightened performance of mf-MELDI-MS for the qualitative analysis of complex mixtures, as targets do not need modification and analysis requires only a few minutes. In addition, GC-MS is suitable for quantitative analysis.

Identification of Verbena officinalis based on ITS sequence analysis and RAPD-derived molecular markers.

Verbenae herba is a widely used drug and consists of the aerial parts of Verbena officinalis (Verbenaceae). Until now, the identification has been performed based on morphological and phytochemical analyses, which are not reliable enough to distinguish Verbena officinalis from other relevant species of the genus Verbena. Hence, impurities and adulterants, negatively influencing the therapeutic effect of the drug, may remain undetected. In an attempt to generate an accurate authentication method we used two different DNA-based approaches: comparison of ITS sequences and molecular markers (RAPD). Both approaches generally enabled discrimination of V. officinalis from the rest of the genus despite the intraspecific variation existing within V. officinalis. The application of the two independent methods, supporting each other, increases the security of identification. For better reproducibility and faster analysis, however, a SCAR marker and primers for HRM were derived from the RAPD results. The SCAR marker could distinguish V. officinalis from all other verbena species except its closest relative V. hastata, while discrimination of V. officinalis even from V. hastata was unproblematic with HRM.

Microarray analysis of host gene expression for comparison between naïve and HSV-1 latent rabbit trigeminal ganglia.

PURPOSE: To analyze the rabbit host global gene expression patterns in uninfected and herpes simplex virus type 1 (HSV-1) latent trigeminal ganglia (TG) for identification of host response-initiated transcriptional changes during the maintenance of viral latency. METHODS: The corneas of eight-week-old New Zealand White rabbits were scarified and inoculated with HSV-1 strain McKrae, 5x10(5) plaque forming units/eye. Corneal infection was verified by slit-lamp examination. Prior to sacrifice at 30 days post infection, ocular swabs confirmed no infectious virus was present. TG were aseptically removed from rabbits and placed in RNA stabilization solution. Host RNA was isolated from two groups of TG, uninfected and HSV-1 latent infected, and used to create labeled cRNA. Labeled cRNA was hybridized to two new and novel custom oligonucleotide rabbit arrays, containing a total of 3,123 probes for rabbit genes. RESULTS: The rabbit TG expressed approximately 80% of genes out of a total of 3,123. A one-way ANOVA performed on the log2 transformed signal ratios showed 611 genes were significantly altered (p< or =0.05) in HSV-1 latent TG. These genes, if annotated, were separated by biologic process categories. Five broad categories were most heavily represented: protein processing, carbohydrate processing, cell adhesion, apoptosis, and host defense and immune response. Sixty of the significantly altered genes were found to be altered by more than 2 fold, and five were altered by more than 4 fold. The genes altered by more than 4 fold were all upregulated and related to host defense and immune response. Viral latency had a large effect on protein processing. Of the differentially expressed genes with an assigned biologic process, 90/349 (25.7%) were associated with protein processing. The next most populated categories were carbohydrate metabolism 39/349 (11.1%) and host defense and immune response 17/349 (4.9%). CONCLUSIONS: The results of this microarray study demonstrate that host gene expression is altered in the HSV-1 latent rabbit TG. The shift in molecular processes at a pathway level reveals the presence of potential therapeutic significance inherent in the maintenance of HSV-1 latency. This is the first large-scale rabbit gene expression study, using microarray analysis, that documents the involvement of host immunity in maintaining HSV-1 latency.

Proanthocyanidins: target compounds as antibacterial agents.

Grape seeds accumulate in huge quantities as byproduct during wine production and are therefore a cheap source for pharmacologically active agents. However, studies prove poor antibacterial activity, and results of analyses are sometimes contradictory. The aim of this study was, thus, to determine the antibacterial activity of grape seed extracts with special focus on the chromatographic characterization of active fractions. In the course of these investigations, extraction protocols were optimized so that microwave-assisted extraction (MAE) guaranteed highest preconcentration efficiency. Proanthocyanidins, monomeric flavonoid aglycones, as well as some of their glycosides could be identified within yielded extracts via high-performance liquid chromatography-mass spectrometry (HPLC-MS). By that means the coherence number of possible isomers of procyanidins was approximated by a newly developed equation. As far as antibacterial activity determined via screening tests is concerned, the extracts generally have been found to be positively responsive toward 10 different gram-positive and gram-negative bacteria strains. After fractionation of the raw extracts, proanthocyanidins P2, P3, P4 and gallate esters P2G and P3G (P = proanthocyanidin consisting of catechin and epicatechin units, n = oligomerization degree, G = gallate ester) were determined as active antibacterial agents toward 10 different pathogens. Only moderate activity was found for monomeric flavonoid fractions.

Quality assessment and quantitative analysis of flavonoids from tea samples of different origins by HPLC-DAD-ESI-MS.

Components of green tea ( Camellia sinensis) have been of considerable interest in recent years because of their potential utility as pharmaceutical agents, particularly for their antioxidant and anticarcinogenic activity. Responding to the increasing scientific validation of numerous health benefits of tea, a comprehensive approach was adopted to carry out analysis for the quality assessment of flavonoids in tea samples of different origins. For this purpose, extraction, separation, and mass spectrometric parameters were optimized. Extraction methods evaluated include reflux extraction, a modified accelerated solvent extraction (ASE), namely, Aquasolv extraction, and microwave-assisted extraction (MAE) using different percentages of solvents. Separation was performed by a specifically developed reversed phase high-performance liquid chromatography (RP-HPLC) method using different C18 and C8 stationary phases. Optimization of extraction techniques clearly proved the performance of MAE, which delivered highest yields in a very short time. Additionally, the comparison with Aquasolv extraction provided new insights, as variations in quantified amounts of target compounds between the extracts could be explained on the basis of thermal degradation and epimerization phenomena. Especially the epimerization phenomenon for catechin/epicatechin oligomers, that is, of procyanidins P 2 and P 3, was observed for the first time. Finally, an optimized extraction and separation system was used for qualitative and quantitative investigations of compounds from different green tea samples from Ceylon (cultivated under biologically controlled conditions), Japan, India, and China as well as from one black tea sample from India.

Identification of carbohydrates by matrix-free material-enhanced laser desorption/ionisation mass spectrometry.

Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) is a sensitive mass spectrometric technique which utilises acidic materials as matrices for laser energy absorption, desorption and ionisation of analytes. These matrix materials produce background signals particularly in the low-mass range and make the detection and identification of small molecules difficult and nearly impossible. To overcome this problem this paper introduces matrix-free material-enhanced laser desorption/ionisation mass spectrometry (mf-MELDI-MS) for the screening and analysis of small molecules such as carbohydrates. For this purpose, 4,4'-azo-dianiline was immobilised on silica gel enabling the absorption of laser energy sufficient for successful desorption and ionisation of low molecular weight compounds. The particle and pore sizes, the solvent system for suspension and the sample preparation procedures have been optimised. The newly synthesised MELDI material delivered excellent spectra with regard to signal-to-noise ratio and detection sensitivity. Finally, wheat straw degradation products and Salix alba L. plant extracts were analysed proving the high performance and excellent behaviour of the introduced material.

Using luminescent nanoparticles as staining probes for Affymetrix GeneChips.

Microarray technology provides efficient access to genetic information using miniaturized, high-density arrays of DNA probes. We investigated the application of luminescent nanoparticles as probes for Affymetrix GeneChips detection without the need for signal amplification. Our goal is to investigate the feasibility of using luminescent nanoparticles as probes in a commercial microarray system without changing its configurations. With the present imaging modality and existing optical excitation and detection systems of the Affymetrix GeneChips, our early results indicate that nanoparticles not only can be used for GeneChip labeling but also are superior to the traditional fluorescent protein streptavidin-phycoerythrin (SAPE). The advantage of the particles lies in a simplified staining procedure, higher photobleaching threshold, and enhanced luminescence signal. The nanoparticles can be used for detection of low-abundance targets without any amplification step. A concentration detection limit of 50 fM has been achieved. This work demonstrates the feasibility of using luminescent nanoparticles as probes for commercial microarray systems, making them less costly, more reproducible, and potentially quantitative.