Antioxidant power measurement in platelet concentrates treated by two pathogen inactivation systems in different blood centres.

BACKGROUND AND OBJECTIVES: The antioxidant power measurement can be useful to validate the execution of the pathogen inactivation treatment of platelet concentrates. The aim of this study is to evaluate the technology on different blood preparations including INTERCEPT and Mirasol treatments that are in routine use in Belgium and Luxemburg. MATERIALS AND METHODS: The antioxidant power measurement was tested on 78 apheresis platelet concentrates and 54 pools of buffy-coats-derived platelet concentrates before and after INTERCEPT treatment. In addition, 100 Reveos platelet pools were tested before and after Mirasol treatment. The antioxidant power was quantified electrochemically using disposable devices and was expressed as equivalent ascorbic acid concentration. RESULTS: Mean results for apheresis platelet concentrates were of 90 ± 14 and 35 ± 10 µmol/l eq. ascorbic acid before and after INTERCEPT treatment, respectively. The mean results for pools of buffy-coats-derived platelet concentrates were of 81 ± 10 and 29 ± 4 eq. µmol/l ascorbic acid before and after INTERCEPT treatment, respectively. For buffy-coats-derived platelet concentrates treated by Mirasol technology, the mean results were of 98 ± 11 and 32 ± 10 µmol/l eq. ascorbic acid before and after illumination, respectively. CONCLUSION: The antioxidant power significantly decreases with pathogen inactivation treatments for platelet concentrates treated by INTERCEPT or Mirasol technologies.

Redox Proteomics and Platelet Activation: Understanding the Redox Proteome to Improve Platelet Quality for Transfusion.

Blood banks use pathogen inactivation (PI) technologies to increase the safety of platelet concentrates (PCs). The characteristics of PI-treated PCs slightly differ from those of untreated PCs, but the underlying reasons are not well understood. One possible cause is the generation of oxidative stress during the PI process. This is of great interest since reactive oxygen species (ROS) act as second messengers in platelet functions. Furthermore, there are links between protein oxidation and phosphorylation, another mechanism that is critical for cell regulation. Current research efforts focus on understanding the underlying mechanisms and identifying new target proteins. Proteomics technologies represent powerful tools for investigating signaling pathways involving ROS and post-translational modifications such as phosphorylation, while quantitative techniques enable the comparison of the platelet resting state versus the stimulated state. In particular, redox cysteine is a key player in platelet activation upon stimulation by different agonists. This review highlights the experiments that have provided insights into the roles of ROS in platelet function and the implications for platelet transfusion, and potentially in diseases such as inflammation and platelet hyperactivity. The review also describes the implication of redox mechanism in platelet storage considerations.

Antioxidant power as a quality control marker for completeness of amotosalen and ultraviolet A photochemical treatments in platelet concentrates and plasma units.

BACKGROUND: Pathogen inactivation treatments such as INTERCEPT aim to make sure blood and blood-derived products are free of pathogens before using them for transfusion purposes. At present, there is no established quality control assay that assesses the completeness of the treatment. As INTERCEPT is a photochemical treatment known to generate reactive oxygen species we sought to use the antioxidant power (AOP) of the blood product as a marker of treatment execution. In this perspective, we evaluated an electrochemically based miniaturized system, the EDEL technology, for measuring the AOP in both platelet concentrates (PCs) and plasma. STUDY DESIGN AND METHODS: Aliquots were withdrawn from PCs or plasma units before and after INTERCEPT treatment and a few microliters were directly deposited into the EDEL sensor for the AOP measurement. The result is expressed in EDEL, an arbitrary unit (micromolar equivalent of ascorbic acid). RESULTS: The INTERCEPT treatment resulted in a significant decrease of the AOP. An AOP threshold of 66.5, 89.0, 59.8, and 131.5 EDEL was determined for apheresis PCs collected from female and male donors, buffy coat PCs, and plasma units, respectively. Below the threshold value, INTERCEPT treatment is considered to be executed. Additionally, we showed that the presence of the photosensitizer in combination with the ultraviolet A illumination is required to observe the AOP decrease. CONCLUSION: The measurement of the AOP of PCs and plasma units can be used to document the completeness of the INTERCEPT treatment.

In vitro study of platelet function confirms the contribution of the ultraviolet B (UVB) radiation in the lesions observed in riboflavin/UVB-treated platelet concentrates.

BACKGROUND: Platelet inactivation technologies (PITs) have been shown to increase platelet storage lesions (PSLs). This study investigates amotosalen/ultraviolet (UV)A- and riboflavin/UVB-induced platelet (PLT) lesions in vitro. Particular attention is given to the effect of UVB alone on PLTs. STUDY DESIGN AND METHODS: Buffy coat-derived PLT concentrates (PCs) were treated with amotosalen/UVA, riboflavin/UVB, or UVB alone and compared to untreated PCs throughout storage. In vitro PLT function was assessed by blood gas and metabolite analyses, flow cytometry-based assays (CD62P, JC-1, annexin V, PAC-1), hypotonic shock response, and static adhesion to fibrinogen-coated wells. RESULTS: In our experimental conditions, riboflavin/UVB-treated PCs showed the most pronounced differences compared to untreated and amotosalen/UVA-treated PCs. The riboflavin/UVB treatment led to a significant increase of anaerobic glycolysis rate despite functional mitochondria, a significant increase of CD62P on Day 2, and a decrease of JC-1 aggregates and increase of annexin V on Day 7. The expression of active GPIIbIIIa (PAC-1) and the adhesion to fibrinogen was significantly increased from Day 2 of storage in riboflavin/UVB-treated PCs. Importantly, we showed that these lesions were caused by the UVB radiation alone, independently of the presence of riboflavin. CONCLUSION: The amotosalen/UVA-treated PCs confirmed previously published results with a slight increase of PSLs compared to untreated PCs. Riboflavin/UVB-treated PCs present significant in vitro PSLs compared to untreated PCs. These lesions are caused by the UVB radiation alone and probably involve the generation of reactive oxygen species. The impact of these observations on clinical use must be investigated.

Extracellular matrix secretion by cardiac fibroblasts: role of microRNA-29b and microRNA-30c.

RATIONALE: MicroRNAs (miRNAs), in particular miR-29b and miR-30c, have been implicated as important regulators of cardiac fibrosis. OBJECTIVE: To perform a proteomics comparison of miRNA effects on extracellular matrix secretion by cardiac fibroblasts. METHODS AND RESULTS: Mouse cardiac fibroblasts were transfected with pre-/anti-miR of miR-29b and miR-30c, and their conditioned medium was analyzed by mass spectrometry. miR-29b targeted a cadre of proteins involved in fibrosis, including multiple collagens, matrix metalloproteinases, and leukemia inhibitory factor, insulin-like growth factor 1, and pentraxin 3, 3 predicted targets of miR-29b. miR-29b also attenuated the cardiac fibroblast response to transforming growth factor-ß. In contrast, miR-30c had little effect on extracellular matrix production but opposite effects regarding leukemia inhibitory factor and insulin-like growth factor 1. Both miRNAs indirectly affected cardiac myocytes. On transfection with pre-miR-29b, the conditioned medium of cardiac fibroblasts lost its ability to support adhesion of rat ventricular myocytes and led to a significant reduction of cardiac myocyte proteins (α-actinin, cardiac myosin-binding protein C, and cardiac troponin I). Similarly, cardiomyocytes derived from mouse embryonic stem cells atrophied under pre-miR-29 conditioned medium, whereas pre-miR-30c conditioned medium had a prohypertrophic effect. Levels of miR-29a, miR-29c, and miR-30c, but not miR-29b, were significantly reduced in a mouse model of pathological but not physiological hypertrophy. Treatment with antagomiRs to miR-29b induced excess fibrosis after aortic constriction without overt deterioration in cardiac function. CONCLUSIONS: Our proteomic analysis revealed novel molecular targets of miRNAs that are linked to a fibrogenic cardiac phenotype. Such comprehensive screening methods are essential to define the concerted actions of miRNAs in cardiovascular disease.

Lysis-on-chip of single target cells following forced interaction with CTLs or NK cells on a dielectrophoresis-based array.

Guiding the interaction of single cells acting as partners in heterotypic interactions (e.g., effectors and targets of immune lysis) and monitoring the outcome of these interactions are regarded as crucial biomedical achievements. In this study, taking advantage of a dielectrophoresis (DEP)-based Laboratory-on-a-chip platform (the DEPArray), we show that it is possible to generate closed DEP cages entrapping CTLs and NK cells as either single cells or clusters; reversibly immobilize a single virus-presenting or tumor cell within the chip at a selected position; move cages and their content to predetermined spatial coordinates by software-guided routing; force a cytotoxic effector to physically interact with a putative target within a secluded area by merging their respective cages; generate cages containing effector and target cells at predetermined E:T ratios; accurately assess cytotoxicity by real-time quantitation of the release kinetics of the fluorescent dye calcein from target cells (>50 lytic events may be tested simultaneously); estimate end points of calcein release within 16 min of initial E:T cell contact; simultaneously deliver Ab-based phenotyping and on-chip lysis assessment; and identify lytic and nonlytic E:T combinations and discriminate nonlytic effector phenotypes from target refractoriness to immune lysis. The proof of principle is provided that DEPArray technology, previously used to levitate and move single cells, can be used to identify highly lytic antiviral CTLs and tumor cells that are particularly refractory to NK cell lysis. These findings are of primary interest in targeted immunotherapy.

Programmable interactions of functionalized single bioparticles in a dielectrophoresis-based microarray chip.

Manipulating single biological objects is a major unmet challenge of biomedicine. Herein, we describe a lab-on-a-chip platform based on dielectrophoresis (DEP). The DEParray is a prototypal version consisting of 320 × 320 arrayed electrodes generating >10,000 spherical DEP cages. It allows the capture and software-guided movement to predetermined spatial coordinates of single biological objects. With the DEParray we demonstrate (a) forced interaction between a single, preselected target cell and a programmable number of either microspheres or natural killer (NK) cells, (b) on-chip immunophenotypic discrimination of individual cells based on differential rosetting with microspheres functionalized with monoclonal antibodies to an inhibitory NK cell ligand (HLA-G), (c) on-chip, real-time (few minutes) assessment of immune lysis by either visual inspection or semiautomated, time-lapse reading of a fluorescent dye released from NK cell-sensitive targets, and (d) manipulation and immunophenotyping with limiting amounts (about 500) cells. To our knowledge, this is the first report describing a DEP-based lab-on-a-chip platform for the quick, arrayed, software-guided binding of individually moved biological objects, the targeting of single cells with microspheres, and the real-time characterization of immunophenotypes. The DEParray candidates as a discovery tool for novel cell:cell interactions with no prior (immuno)phenotypic knowledge.

Sulfenylome analysis of pathogen-inactivated platelets reveals the presence of cysteine oxidation in integrin signaling pathway and cytoskeleton regulation.

Essentials Cysteine oxidation to sulfenic acid plays a key role in redox regulation and signal transduction. Platelet sulfenylome was studied by quantitative proteomics in pathogen inactivated platelets. One hundred and seventy-four sulfenylated proteins were identified in resting platelets. Pathogen inactivation oxidized integrin ßIII, which could activate the mitogen-activated protein kinases pathway. ABSTRACT: Background Cysteine-containing protein modifications are involved in numerous biological processes such redox regulation or signal transduction. During the preparation and storage of platelet concentrates, cell functions and protein regulations are impacted. In spite of several proteomic investigations, the platelet sulfenylome, ie, the proteins containing cysteine residues (R-SH) oxidized to sulfenic acid (R-SOH), has not been characterized. Methods A dimedone-based sulfenic acid tagging and enrichment coupled to a mass spectrometry identification workflow was developed to identify and quantify the sulfenic acid-containing proteins in platelet concentrates treated or not with an amotosalen/ultraviolet A (UVA) pathogen inactivation technique. Results One hundred and seventy-four sulfenylated proteins were identified belonging mainly to the integrin signal pathway and cytoskeletal regulation by Rho GTPase. The impact on pathogen inactivated platelet concentrates was weak compared to untreated ones where three sulfenylated proteins (myosin heavy chain 9, integrin ßIII, and transgelin 2) were significantly affected by amotosalen/UVA treatment. Of particular interest, the reported oxidation of cysteine residues in integrin ßIII is known to activate the receptor αIIbßIII. Following the pathogen inactivation, it might trigger the phosphorylation of p38MAPK and explain the lesions reported in the literature. Moreover, procaspase activating compound-1 (PAC-1) binding assays on platelet activation showed an increased response to adenosine diphosphate exacerbated by the tagging of proteins with dimedone. This result corroborates the hypothesis of an oxidation-triggered activation of αIIbßIII by the pathogen inactivation treatment. Conclusions The present work completes missing information on the platelet proteome and provides new insights on the effect of pathogen inactivation linked to integrin signaling and cytoskeleton regulation.

Hypoxia and hypocapnia storage of γ-irradiated red cell concentrates.

BACKGROUND: γ-irradiation is used to treat red blood cell (RBC) concentrates (RCCs) transfused to immunosuppressed patients. This treatment damages RBCs and increases storage lesions. Several studies have shown the beneficial effect of reducing O2 content during RBC storage. The present research work investigated the effect of γ-irradiation on RCCs stored under normal and hypoxia/hypocapnia conditions. MATERIALS AND METHODS: O2 concentration (measured as oxyhaemoglobin fraction, sO2) and ABO-matched RCCs from whole blood donations, leukoreduced and prepared in phosphate, adenine, glucose, guanosine, saline and mannitol (PAGGSM) were pooled and split in two identical RCCs within 24 h post donation. One bag (Hx) was submitted to O2 and CO2 adsorption for 3 h on an orbital shaker at 22±2 °C and then transferred to a storage bag impermeable to gas. The other bag (Ctrl) was left as it was. The two bags were then stored at 4 °C. γ-irradiation (25 Gy) was applied at day 2 or 14, and the RCCs were stored until day 43. Different parameters (metabolites, haemolysis, morphology) were measured. RESULTS: Starting sO2 values were 63.7±18.4% (n=12) in Ctrl and 20.8±9.8% (n=12) in Hx bags, and reached 90.8±9.1% and 6.6±5.9% at day 43, respectively. As expected, an increase in glycolysis rate was observed after deoxygenation. Extracellular potassium concentrations were identical and reached around 70 mM at expiry with an irradiation-dependent kinetic release. No difference in haemolysis was observed after irradiation on day 2 in either group (<0.40%, p>0.9999). When irradiated at day 14, haemolysis was lower (p=0.033) in RCCs under hypoxia at the end of storage (day 28, 0.67±0.16%) compared to control (1.06±0.33%). Percentages of spherocytes were lower under hypoxia. DISCUSSION: The storage under hypoxia provided equivalent storage when RCCs were irradiated at day 2 and was advantageous when irradiated at day 14. In summary, O2-depletion of RCCs enable a better storage of RBCs, particularly when late irradiation is applied.

Highlights from the 2010 BAS/BSCR spring meeting: New Frontiers in Cardiovascular Research.

The British Atherosclerosis Society (BAS)/British Society for Cardiovascular Research (BSCR) spring meeting was held in Manchester, UK, on 7-8 June 2010. Experts in the field of systems biology, proteomics, metabolomics and miRNAs presented how these techniques can be used to discover 'New Frontiers in Cardiovascular Research'. The conference was attended by over 150 participants, mainly from the UK. A total of 2 days of presentations and a poster session with 55 posters provided the possibility to discuss the latest research results and showed the opportunities that new techniques can offer in cardiovascular research.

Electrochemical reactions and ionization processes.

Electrochemical or photo-electrochemical reactions in both electrospray ionization and laser desorption ionization are discussed stressing the role of the electrode reaction in influencing the ionization process. In particular, upon application of a high voltage during electrospray ionization, the emitter includes a working electrode, where redox reactions are observed, such as electro-generation of benzoquinone and metal ions. In contrast, the target plate in laser-induced desorption ionization also acts as a photo-electrode, especially when modified with a mesoporous semiconductor. We illustrate here how these electrochemical reactions can be used for tagging purposes, and for oxidative or reductive dissociation reactions.

Oxygen in Red Blood Cell Concentrates: Influence of Donors' Characteristics and Blood Processing.

Objective: Unexpectedly wide distribution (<10 to >90%) of hemoglobin oxygen saturation (sO2) within red cell concentrates (RCCs) has recently been observed. Causes of such variability are not yet completely explained whereas the roles of oxygen and oxidative lesions during the storage of RCCs are known. The objectives of the present study are to characterize sO2 distribution in RCCs produced in a Swiss blood center and to investigate the influence of processing and donors' characteristics. Methods: The level of sO2 was measured in 1701 leukocyte-depleted RCCs derived from whole blood donations in both top-bottom (TB; component filtered, SAGM) and top-top (TT; whole blood filtration, PAGGSM) RCCs. The sO2 value was measured non-invasively through the PVC bag prior to storage by resonance Raman spectroscopy. Gender, age, blood type, hemoglobin level, and living altitude of donors, as well as process method and time-to-process were recorded. Results: Overall, the sO2 exhibited a wide non-Gaussian distribution with a mean of 51.2 ± 18.5%. Use of top-top kits resulted in a 16% higher sO2 (P < 0.0001) than with top-bottom ones. Waiting time before processing only had a modest impact, but the blood processing itself reduced the sO2 by almost 12% (P < 0.0001). sO2 was also significantly affected by some donors' characteristics. RCCs from men exhibited 25% higher sO2 (P < 0.0001) than those donated by women. Multivariate analysis revealed that the apparent correlation observed with hemoglobin level and age was actually due to multicollinearity with the sex variable. Finally, we noticed no significant differences across blood type but found that altitude of residence was associated with the sO2 (i.e., higher in higher living place). Conclusion: These data confirm wide sO2 distribution in RCCs reported recently. The sO2 was impacted by the processing and also by donors' characteristics such as the gender and the living altitude, but not by the hemoglobin level, blood group and donor age. This study provides new hints on the factors influencing red blood cells storage lesions, since they are known to be related to O2 content within the bags, giving clues to better process and to better store RCCs and therefore potentially improve the efficacy of transfusion.

Transfusion-transmitted cytomegalovirus: behaviour of cell-free virus during blood component processing. A study on the safety of labile blood components in Switzerland.

BACKGROUND: Nowadays, most blood products are leukocyte-reduced. After this procedure, the residual risk for transfusion transmitted cytomegalovirus (TT-CMV) is mostly attributed to cell-free viruses in the plasma of blood donors following primary infection or viral reactivation. Here, objectives are: 1) to study the behaviour of cell-free CMV through the blood component processing; 2) to determine the anti-CMV seroprevalence, the level of viremia, the window-period in blood donor population; and 3) to identify cases of TT-CMV in bone marrow transplant (BMT) recipients. MATERIALS AND METHODS: Cell-free CMV was injected into blood bags originating from regular donors. Blood components were processed according to either the CompoSelect® or the CompoFlow® (Fresenius Kabi AG) techniques. Samples were analysed at each step for presence of virus DNA using quantitative polymerase chain reaction (PCR). The anti-CMV seroprevalence in our donor population was taken from our donor data system. The viremia was assessed in pooled plasmas samples from routine donations by quantitative PCR. Medical charts of 165 BMT anti-CMV seronegative recipients/anti-CMV seronegative donors who received CMV-unscreened blood products were reviewed. RESULTS: Cell-free CMV passes without any decrease in viral load through all stages of blood processing. The anti-CMV seroprevalence was 46.13%. Four DNA positive samples out of 42,240 individual blood donations were identified (0.009%); all had low levels of viremia (range 11-255 IU/mL). No window-period donation was identified. No TT-CMV was found. DISCUSSION: Cell-free CMV remains a concern with current blood component processing as it passes through all the processes. However, since low levels of CMV DNA were identified in the donations tested, and no BMT recipients had TT-CMV, the residual threat of TT-CMV after leukocyte reduction appears to be very low.

Sandwich mixer-reactor: influence of the diffusion coefficient and flow rate ratios.

A sandwich mixer consists of mixing two solutions in a channel, one central laminar flow being sandwiched between two outer flow solutions. The present numerical study considers the convection-diffusion of two reacting species A and B, provided respectively by the two incoming solutions. The simulations show how the diffusion coefficient, flow rate and species concentration ratios influence, via the transversal diffusion length and reaction kinetics, the reaction extent at the end of the sandwich mixer. First, this extent can be enhanced up to 60% if the species with the lowest diffusion coefficient is located in the outer solutions where the flow velocity is small compared to that of the central part (higher residence time). Secondly, decreasing the outer flow rates (to confine the reaction close to the walls) and increasing the local concentration to keep the same flux ratio improve the extent by 300%. Comparison with a bi-lamination passive mixer, with an ideal mixer and an electro-osmotic driven flow mixer is presented. These conclusions are also demonstrated for consecutive reactions, showing an amplification of the effects described above. The results are also presented versus the residence time in the mixer-reactor to show the time window for which the gain is appreciable.

Magnetic forces produced by rectangular permanent magnets in static microsystems.

Finite element numerical simulations were carried out in 2D geometries to map the magnetic field and force distribution produced by rectangular permanent magnets as a function of their size and position with respect to a microchannel. A single magnet, two magnets placed in attraction and in repulsion have been considered. The goal of this work is to show where magnetic beads are preferentially captured in a microchannel. These simulations were qualitatively corroborated, in one geometrical case, by microscopic visualizations of magnetic bead plug formation in a capillary. The results show that the number of plugs is configuration dependent with: in attraction, one plug in the middle of the magnets; in repulsion, two plugs near the edges of the magnets; and with a single magnet, a plug close to the center of the magnet. The geometry of the magnets (h and l are the height and length of the magnets respectively) and their relative spacing s has a significant impact on the magnetic flux density. Its value inside a magnet increases with the h/l ratio. Consequently, bar magnets produce larger and more uniform values than flat magnets. The l/s ratio also influences the magnetic force value in the microchannel, both increasing concomitantly for all the configurations. In addition, a zero force zone in the middle appears in the attraction configuration as the l/s ratio increases, while with a single magnet, the number of maxima and minima goes from one to two, producing two focusing zones instead of only one.

Magnetic track array for efficient bead capture in microchannels.

Magnetism-based microsystems, as those dedicated to immunoaffinity separations or (bio)chemical reactions, take benefit of the large surface area-to-volume ratio provided by the immobilized magnetic beads, thus increasing the sensitivity of the analysis. As the sensitivity is directly linked to the efficiency of the magnetic bead capture, this paper presents a simple method to enhance the capture in a microchannel. Considering a microchannel surrounded by two rectangular permanent magnets of different length (L (m) = 2, 5, 10 mm) placed in attraction, it is shown that the amount of trapped beads is limited by the magnetic forces mainly located at the magnet edges. To overcome this limitation, a polyethylene terephthalate (PET) microchip with an integrated magnetic track array has been prototyped by laser photo-ablation. The magnetic force is therefore distributed all along the magnet length. It results in a multi-plug bead capture, observed by microscope imaging, with a magnetic force value locally enhanced. The relative amount of beads, and so the specific binding surface for further immunoassays, presents a significant increase of 300% for the largest magnets. The influence of the track geometry and relative permeability on the magnetic force was studied by numerical simulations, for the microchip operating with 2-mm-long magnets.

Electrospray micromixer chip for on-line derivatization and kinetic studies.

An electrospray microchip for mass spectrometry comprising an integrated passive mixer to carry out on-chip chemical derivatizations is described. The microchip fabricated using UV-photoablation is composed of two microchannels linked together by a liquid junction. Downstream of this liquid junction, a mixing unit made of parallel oblique grooves is integrated to the microchannel in order to create flow perturbations. Several mixer designs are evaluated. The mixer efficiency is investigated both by fluorescence study and mass spectrometric monitoring of the tagging reaction of cysteinyl peptides with 1,4-benzoquinone. The comparisons with a microchip without a mixing unit and a kinetic model are used to assess the efficiency of the mixer showing tagging kinetics close to that of bulk reactions in an ideally mixed reactor. As an ultimate application, the electrospray micromixer is implemented in a LC-MS workflow. On-line derivatization of albumin tryptic peptides after a reversed-phase separation and counting of their cysteines drastically enhance the protein identification.

General overview of blood products in vitro quality: Processing and storage lesions.

Blood products are issued from blood collection. Collected blood is immediately mixed with anticoagulant solutions that immediately induce chemical and/or biochemical modifications. Collected blood is then transformed into different blood products according to various steps of fabrication. All these steps induce either reversible or irreversible "preparation-related" lesions that combine with "storage-related" lesions. This short paper aims to provide an overview of the alterations that are induced by the "non-physiological" processes used to prepare blood products that are used in clinical practice.

Electronic sorting and recovery of single live cells from microlitre sized samples.

Sorting and recovering specific live cells from samples containing less than a few thousand cells have become major hurdles in rare cell exploration such as stem cell research, cell therapy and cell based diagnostics. We describe here a new technology based on a microelectronic chip integrating an array of over 100,000 independent electrodes and sensors which allow individual and parallel single cell manipulation of up to 10,000 cells while maintaining viability and proliferation capabilities. Manipulation is carried out using dynamic dielectrophoretic traps controlled by an electronic interface. We also demonstrate the capabilities of the chip by sorting and recovering individual live fluorescent cells from an unlabeled population.

Multitrack electrospray chips.

Multitrack electrospray chips (MTEC) were fabricated by UV-photoablation of polyethylene terephthalate (PET) substrates. They are composed of an array of up to six microchannels that are successively used as electrospray ionization (ESI) emitters for mass spectrometry (MS). There is no requirement for alignment of the different spraying microchannels with the mass spectrometer orifice. The MTEC is thus fixed in front of the mass spectrometer and the successive MS analyses are performed without moving the chip. The sequential electrospraying by successive application of an identical high voltage in each off-axis microchannel was evaluated for the fast screening of peptides and proteins. The counting of cysteines in peptides through chemical modification and the relative quantification of a peptide in two samples are presented herein as two original strategies based on this new analytical tool.