40 W of supercontinuum generated by a self-pulsed pump-sharing oscillator-amplifier.

We demonstrate an all-fiber supercontinuum (SC) source delivering up to 40 W of average power ranging from 750 to 2200 nm. The laser source is based on a self-Q-switched pump-sharing oscillator-amplifier. The self-Q-switched master oscillator generates giant pulses, amplified in the high-power stage. Finally, a passive fiber acts as a nonlinear stage, improving the spectrum flatness as well as the spectral broadening. To the best of our knowledge, this is the first time that a pump-sharing oscillator-amplifier is used for SC generation and is based on the use of a submeter Ytterbium-doped fiber length inside the oscillator.

Photoacclimation of the polar diatom Chaetoceros neogracilis at low temperature.

Polar microalgae face two major challenges: 1- growing at temperatures (-1.7 to 5°C) that limit enzyme kinetics; and 2- surviving and exploiting a wide range of irradiance. The objective of this study is to understand the adaptation of an Arctic diatom to its environment by studying its ability to acclimate to changes in light and temperature. We acclimated the polar diatom Chaetoceros neogracilis to various light levels at two different temperatures and studied its growth and photosynthetic properties using semi-continuous cultures. Rubisco content was high, to compensate for low catalytic rates, but did not change detectably with growth temperature. Contrary to what is observed in temperate species, in C. neogracilis, carbon fixation rate (20 min 14C incorporation) equaled net growth rate (µ) suggesting very low or very rapid (<20 min) re-oxidation of the newly fixed carbon. The comparison of saturation irradiances for electron transport, oxygen net production and carbon fixation revealed alternative electron pathways that could provide energy and reducing power to the cell without consuming organic carbon which is a very limiting product at low temperatures. High protein contents, low re-oxidation of newly fixed carbon and the use of electron pathways alternative to carbon fixation may be important characteristics allowing efficient growth under those extreme environmental conditions.

Design, Synthesis and SAR in 2,4,7-Trisubstituted Pyrido[3,2-d]Pyrimidine Series as Novel PI3K/mTOR Inhibitors.

This work describes the synthesis, enzymatic activities on PI3K and mTOR, in silico docking and cellular activities of various uncommon 2,4,7 trisubstituted pyrido[3,2-d]pyrimidines. The series synthesized offers a chemical diversity in C-7 whereas C-2 (3-hydroxyphenyl) and C-4 groups (morpholine) remain unchanged, in order to provide a better understanding of the molecular determinants of PI3K selectivity or dual activity on PI3K and mTOR. Some C-7 substituents were shown to improve the efficiency on kinases compared to the 2,4-di-substituted pyrimidopyrimidine derivatives used as references. Six novel derivatives possess IC50 values on PI3Kα between 3 and 10 nM. The compounds with the best efficiencies on PI3K and mTOR induced micromolar cytotoxicity on cancer cell lines possessing an overactivated PI3K pathway.

30 TW and 33 fs pulses delivered by a Ti:Sa amplifier system seeded with a frequency-doubled fiber laser.

We report a full experimental comparison study on the injection of a Ti:Sa multi-terawatt amplifier chain with a standard 15 fs Ti:Sa oscillator and 35 fs frequency-doubled fiber oscillator. The study highlights that the Ti:Sa oscillator, with high performance in terms of pulse duration and spectral width, can be replaced by the frequency-doubled fiber oscillator to seed Ti:Sa amplifier chains almost without any compromise on the output pulse duration and picosecond contrast. Finally, we demonstrate for the first time to our knowledge a 30 TW and 33 fs Ti:Sa amplifier injected by a fiber oscillator.

Capillary electrophoresis with dual detection UV/C4D for monitoring myrosinase-mediated hydrolysis of thiol glucosinolate designed for gold nanoparticle conjugation.

Capillary electrophoresis (CE) with dual UV and conductivity detection was used for the first time to monitor the functionalization of gold nanoparticles (AuNPs), a process catalyzed by an enzyme, myrosinase (Myr). A thiol glucosinolate (GL-SH) designed by our group was used as substrate. Hydrolysis of free and immobilized GL-SH was characterized using off-line and on-line CE-based enzymatic assays. The developed approaches were validated using sinigrin, a well-referenced substrate of Myr. Michaelis-Menten constant of the synthetized GL-SH was comparable to sinigrin, showing that they both have similar affinity towards Myr. It was demonstrated that transverse diffusion of laminar flow profiles was well adapted for in-capillary Mixing of nanoparticles (AuNPs) with proteins (Myr) provided that the incubation time is inferior to 20 min. Only low reaction volume (nL to few µL) and short analysis time (<5 min) were required. The electrophoretic conditions were optimized in order to evaluate and to confirm the AuNPs stability before and after functionalization by CE/UV based on surface plasmon resonance band red-shifting. The hydrolysis of the functionalized AuNPs was subsequently evaluated using the developed CE-C4D/UV approach. Repeatabilities of enzymatic assays, of electrophoretic analyses and of batch-to-batch functionalized AuNPs were excellent.

Effect of modified di- and trisaccharides on hyaluronidase activity assessed by capillary electrophoresis-based enzymatic assay.

The activity of eukaryote hydrolase-type of hyaluronidases was studied using a miniaturized capillary electrophoresis (CE) assay developed in our laboratory. Few nanoliters of reagents are sufficient and no labeling is required for this assay. The effect of natural and original synthetic effectors of hyaluronidase was evaluated. These di- and trisaccharides from linkage region of proteoglycans were synthesized in 30-40 steps from monomeric units using classical protection, deprotection, glycosylation and deoxygenation reactions. The influence of the chain length (di/trisaccharide), the modification type (methoxy/deoxy) and its position (2/4/6) was studied. The inhibition and/or activation percentages were determined at two concentrations of effectors; 0.2 mM and 2 mM. The half maximal effective concentration (EC50) values were evaluated (n = 2) for the most effective inhibitors (∼1 mM) and activators (∼0.2 mM). Results showed that hyaluronidase was mostly inhibited in a concentration-dependent fashion by a deoxy modification and activated by a methoxy modification. Trisaccharides were found to be more effective on hyaluronidase activity than disaccharides. Position 4 was found to be more favorable for hyaluronidase activity than position 6 and the activity in position 2 was negligible. For a better understanding of the enzyme function mode, the inhibition constant (Ki) was also evaluated by CE (Ki ∼ 2 mM). These results are of great interest especially as few activators of hyaluronidase are presented in the literature.

Monitoring of successive phosphorylations of thymidine using free and immobilized human nucleoside/nucleotide kinases by Flow Injection Analysis with High-Resolution Mass Spectrometry.

Nucleosides and their analogues play a crucial role in the treatment of several diseases including cancers and viral infections. Their therapeutic efficiency depends on their capacity to be converted to the active nucleoside triphosphates form through successive phosphorylation steps catalyzed by nucleoside/nucleotide kinases. It is thus mandatory to develop an easy, rapid, reliable and sensitive enzyme activity tests. In this study, we monitored the three-step phosphorylation of thymidine to thymidine triphosphate respectively by (1) human thymidine kinase 1 (hTK1), (2) human thymidylate kinase (hTMPK) and (3) human nucleoside diphosphate kinase (hNDPK). Free and immobilized kinase activities were characterized by using the Michaelis-Menten kinetic model. Flow Injection Analysis (FIA) with High-Resolution Mass Spectrometry (HRMS) was used as well as capillary electrophoresis (CE) with UV detection. The three-step cascade phosphorylation of thymidine was also monitored. FIA-HRMS allows a sensitive and rapid evaluation of the phosphorylation process. This study proposes simple, rapid, efficient and sensitive methods for enzyme kinetic studies and successive phosphorylation monitoring with immobilized enzymes.

µJ-level Raman-assisted fiber optical parametric chirped-pulse amplification.

We experimentally demonstrate the amplification of chirped pulses in a fiber optical parametric chirped pulse amplifier up to 1 µJ. This high energy level originates from combined Raman and parametric processes in a specially designed solid core photonic bandgap fiber. Output pulses are recompressed up to 560 fs. These performances make this all-fiber system compatible with first stages of bulk amplification chains.

Simultaneous elastase-, hyaluronidase- and collagenase-capillary electrophoresis based assay. Application to evaluate the bioactivity of the red alga Jania rubens.

There have been many efforts to search for affordable and efficient cosmetic ingredients from natural sources and to evaluate their bioactivities using eco-responsible tools. Hyaluronidase, elastase and collagenase are responsible for the degradation of the main components of the extracellular matrix, namely the hyaluronic acid, elastin and collagen, respectively. The aim of this work was to develop a single capillary electrophoresis method to monitor simultaneously the activities of these three enzymes, without reactant immobilization or radioactivity use. The developed approach was used to evaluate the bioactivity of the red alga Jania rubens after microwave- or electrochemical-assisted extraction. For this purpose, the incubation time, the reactant concentrations, the separation buffer and the detection system were carefully chosen. CE with double detection system, LIF and HRMS connected in series, was used to ensure the simultaneous analysis of the substrates and products of the three enzymatic reactions. The optimized enzymatic conditions allowed the use of the same protocol to assess the 3 enzyme activities. These conditions consisted of 10 min pre-incubation of the enzyme (with alga extract) at 37 °C; 10 min incubation with the substrate at 37 °C and 10 min stop-time at 90 °C. 1.4 nL of each reaction mixture were co-injected into a 85 cm total length capillary using short-end injection. Ammonium acetate (50 mM, pH 9.0) was used for electrophoretic separation. All substrates and products were simultaneously detected in less than 10 min with good peak symmetry and efficiency, sufficient intra-day and inter-day repeatabilities (RSD < 4.5%; n = 3) and excellent LOQ (<5 nM). The results obtained using this multiple CE-based enzymatic assay showed the significant effect of Jania rubens ethanolic extracts on elastase, hyaluronidase and the metalloproteinase MMP-1.

Use of chromatographic and electrophoretic tools for assaying elastase, collagenase, hyaluronidase, and tyrosinase activity.

Elastase, collagenase, hyaluronidase and tyrosinase, are very interesting enzymes due to their direct implication in skin aging and as therapeutic hits. Different techniques can be used to study these enzymes and to evaluate the influence of effectors on their kinetics. Nowadays, analytical techniques have become frequently used tools for miniaturizing enzyme assays. The main intention of this article is to review chromatographic and electrophoretic tools that study the four enzymes above mentioned. More specifically, the use of high-performance liquid chromatography and capillary electrophoresis and their derivative techniques for monitoring these enzymes will be investigated. The advantages and limitations of these assays will also be discussed. The original use of microscale thermophoresis and thin layer chromatography in this domain will also be covered.

Macroalga Padina pavonica water extracts obtained by pressurized liquid extraction and microwave-assisted extraction inhibit hyaluronidase activity as shown by capillary electrophoresis.

Hyaluronidase degrades hyaluronic acid, the principal component of the extracellular matrix. Inhibition of this enzyme is thus expected to hinder skin aging. Brown alga Padina pavonica activity toward hyaluronidase was evaluated using capillary electrophoresis (CE)-based enzymatic assays. This green technique allows evaluation of the biological activity of the natural material in an economic manner. Pressurized liquid extraction (PLE), microwave assisted extraction (MAE), supercritical fluid extraction and electroporation extraction techniques were used. Extraction conditions were optimized to obtain cosmetically acceptable Padina pavonica extracts with the best inhibition activity. CE-based assays were conducted using only a few nanoliters of reactants, a capillary of 60cm total length and of 50µm internal diameter, +20kV voltage for separation in 50mM ammonium acetate buffer (pH 9.0) and 200nm wavelength for detection. The reaction mixture was incubated for 1h and CE analysis time was about 11min. A novel online CE-assay using transverse diffusion of laminar flow profiles for in-capillary reactant mixing allowed efficient monitoring of hyaluronidase kinetics with Km and Vmax equal to 0.46±0.04mgmL-1 and 137.1±0.3nMs-1 (r2=0.99; n=3), respectively. These values compared well with literature, which validates the assay. Water extracts obtained by PLE (60°C; 2 cycles) and MAE (60°C; 1000W; 2min) presented the highest anti-hyaluronidase activity. The half maximal effective concentration (IC50) of water PLE extract was 0.04±0.01mgmL-1 (r2=0.99; n=3). This value is comparable to the one obtained for Einsenia bicyclis phlorotannin fractions (IC50=0.03mgmL-1), which makes Padina pavonica bioactivity very promising.

Preliminary recovery study of a commercial molecularly imprinted polymer for the extraction of glyphosate and AMPA in different environmental waters using MS.

A commercial molecularly imprinted polymer (MIP) dedicated to glyphosate (GLY) and its main metabolite, aminomethylphosphonic acid (AMPA), was lately assessed as "POCIS-like" sampler on mineral water. The obtained results were encouraging with 111 and 122 mL day-1 as sampling rates for GLY and AMPA, respectively. Therefore, before applying this passive sampler to environmental waters, the commercial phase was tested on different water matrices as a solid-phase extraction (SPE) device. The SPE protocol was carried on 250 mg of MIP with the following three steps: conditioning by Milli-Q water, loading of the sample (15 mL), and elution of the analytes by 4 mL 0.1 M HCl that were evaporated to dryness and recovered in 15 mL of the suitable solvent for analysis. This protocol was first applied to mineral water spiked by GLY and AMPA at environmental concentration levels (25-750 ng L-1). Analyses were carried out by ultra-performance liquid chromatography hyphenated to tandem mass after derivatization of GLY and AMPA by 9-fluorenylmethylchloroformate. The linear correlation between concentrations measured with and without SPE on MIP was proved.Furthermore, other extractions showed that high concentrations of metal ion interferents (lead(II), cadmium(II), and zinc(II)) in groundwaters did not reduce SPE performance of the MIP.Then, concentration assays were undertaken and brought noteworthy results, such as the recovery of 80% GLY and AMPA from groundwater spiked at 10 ng L-1 and concentrated 100 times. For this purpose, ion exclusion chromatography hyphenated to mass was applied without previous derivatization of the analytes. The same concentration factor and analytical method were applied to 100 ng L-1 spiked sea water with recoveries of 96% for GLY and 121% for AMPA.

Laboratory calibration of a POCIS-like sampler based on molecularly imprinted polymers for glyphosate and AMPA sampling in water.

For more than 15 years, integrative passive sampling has been successfully used for monitoring contaminants in water, but no passive sampling device exists for strongly polar organic compounds, such as glyphosate. We thus propose a polar organic chemical integrative sampler (POCIS)-like tool dedicated to glyphosate and its main degradation product aminomethylphosphonic acid (AMPA), and describe the laboratory calibration of such a tool for calculating the sampling rates of glyphosate and AMPA. This passive sampler consists of a POCIS with molecularly imprinted polymer as a receiving phase and a polyethersulfone diffusion membrane. The calibration experiment for the POCIS was conducted for 35 days in a continuous water-flow-through exposure system. The calibration results show that the sampling rates are 111 and 122 mL day-1 for glyphosate and AMPA respectively, highlighting the potential interest in and the applicability of this method for environmental monitoring. The influence of membrane porosity on the glyphosate sampling rate was also tested. Graphical Abstract ᅟ.

Hyaluronidase reaction kinetics evaluated by capillary electrophoresis with UV and high-resolution mass spectrometry (HRMS) detection.

The biology of hyaluronidase activity on age related turnover of the hyaluronic acid (HA) in skin dermis and epidermis has not been established. Elucidation of this phenomenon enables discovery of novel compounds for skin health. As a simple and green technique, capillary electrophoresis (CE) was used for the first time for the determination of the kinetic constants (Km, Vmax and IC50) of the enzymatic degradation of HA. Reaction products were identified using CE/high-resolution mass spectrometry (HRMS) after appropriate optimization. Best results in terms of signal sensitivity were obtained using 10 mM ammonium acetate (pH 9.0) BGE, a sheath liquid composed of methanol-water (80:20, v/v) with 0.02% (v/v) formic acid at 10 µL min-1 and an ESI voltage at -4 kV. Km and Vmax were determined (n = 3) using CE/UV at 200 nm as 0.24 ± 0.02 mg mL-1 and 150.4 ± 0.1 nM s-1, respectively. They were also successfully obtained by CE/HRMS (n = 3) with Km of 0.49 ± 0.02 mg mL-1 and Vmax of 155.7 ± 0.2 nM s-1. IC50 of a standard natural inhibitor, epigallocatechin gallate, was also determined by CE-UV/HRMS. Kinetic constant values obtained by CE compared well with literature which validated the developed CE-based assay. In addition, the activity of homemade tetrasaccharides of biotinylated chondroitin sulfate CS-A or CS-C (4- or 6- sulfated in a homogeneous or heterogeneous way) on the hydrolysis reaction of hyaluronidase was evaluated. Hyaluronidase was mostly dose-dependently inhibited by CS-A tetrasaccharides sulfated in a homogeneous way. Two trisaccharides from truncated linkage region of proteoglycans were also tested as inhibitors or activators. CE-based assay showed that even a small modification of one hydroxyl group changes the influence on hyaluronidase activity. CE-based assay can be used for the screening of natural and synthetic inhibitors of hyaluronidase activity for cosmetic and therapeutic applications.

Microalgae amino acid extraction and analysis at nanomolar level using electroporation and capillary electrophoresis with laser-induced fluorescence detection.

Amino acids play a key role in food analysis, clinical diagnostics, and biochemical research. Capillary electrophoresis with laser-induced fluorescence detection was used for the analysis of several amino acids. Amino acid labeling with fluorescein isothiocyanate was conducted using microwave-assisted derivatization at 80°C (680 W) during only 150 s. Good electrophoretic resolution was obtained using a background electrolyte composed of sodium tetraborate buffer (100 mM; pH 9.4) and ß-cyclodextrin (10 mM), and the limits of quantification were 3-30 nM. The developed capillary electrophoresis with laser-induced fluorescence method was used to analyze amino acids in Dunaliella salina green algae grown under different conditions. A simple extraction technique based on electroporation of the cell membrane was introduced. A home-made apparatus allowed the application of direct and alternating voltages across the electrochemical compartment containing a suspension of microalgae in distilled water at 2.5 g/L. A direct voltage of 12 V applied for 4 min gave the optimum extraction yield. Results were comparable to those obtained with accelerated-solvent extraction. The efficiency of electroporation in destroying microalgae membranes was shown by examining the algae surface morphology using scanning electron microscopy. Stress conditions were found to induce the production of amino acids in Dunaliella salina cells.

Optical Dark Rogue Wave.

Photonics enables to develop simple lab experiments that mimic water rogue wave generation phenomena, as well as relativistic gravitational effects such as event horizons, gravitational lensing and Hawking radiation. The basis for analog gravity experiments is light propagation through an effective moving medium obtained via the nonlinear response of the material. So far, analogue gravity kinematics was reproduced in scalar optical wave propagation test models. Multimode and spatiotemporal nonlinear interactions exhibit a rich spectrum of excitations, which may substantially expand the range of rogue wave phenomena, and lead to novel space-time analogies, for example with multi-particle interactions. By injecting two colliding and modulated pumps with orthogonal states of polarization in a randomly birefringent telecommunication optical fiber, we provide the first experimental demonstration of an optical dark rogue wave. We also introduce the concept of multi-component analog gravity, whereby localized spatiotemporal horizons are associated with the dark rogue wave solution of the two-component nonlinear Schrödinger system.

Human neutrophil elastase inhibition studied by capillary electrophoresis with laser induced fluorescence detection and microscale thermophoresis.

Capillary electrophoresis-laser induced fluorescence (CZE-LIF) and microscale thermophoresis (MST) were used for the first time to study the inhibition of human neutrophil elastase (HNE). We recently studied HNE kinetics (Km and Vmax) by developing an in-capillary CZE-LIF assay based on transverse diffusion of laminar flow profiles (TDLFP) for reactant mixing. In this work, the former assay was adapted to monitor HNE inhibition. Two natural well known HNE inhibitors from the triterpene family, ursolic acid and oleanolic acid, were tested to validate the developed assay. Since the solubility of pentacyclic triterpenes in aqueous media where the enzymatic reaction will take place is limited, the effect of DMSO and ethanol on HNE was studied using microscale thermophoresis (MST). An agglomeration of the enzyme was revealed when preparing the inhibitor in 5% (v/v) DMSO. This phenomenon did not occur in the presence of ethanol. Therefore, ethanol was used as inhibitor solvent, at a limited percentage of 20% (v/v). In these conditions and after optimization of the TDLFP approach, the repeatability (RSD on migration times and peak-areas inferior to 2.2%) of the CZE-LIF assay and the sensitivity (LOQ of few nM) were found to be satisfactory for conducting inhibition assays. IC50 values for ursolic and oleanolic acid were successfully determined. They were respectively equal to 5.62±0.10µM (r(2)=0.9807; n=3) and to 8.21±0.23µM (r(2)=0.9887; n=3). Excellent agreement was found between the results obtained by CE and those reported in literature which validates the developed method. Particularly, the CE-based assay is able to rank HNE inhibitors relative to each other. Furthermore, MST technique was used for evaluating HNE interaction with the ursolic acid. Up to 16 capillaries were automatically processed to obtain in one titration experiment the dissociation constant for the HNE-ursolic acid complex. Ki was found to be 2.72±0.66µM (n=3) which is in excellent agreement with the value determined by CE enzyme inhibition studies (Ki=2.81µM) confirming the reliability of the developed CE assay and the competitive inhibition mode of ursolic acid.

Assaying human neutrophil elastase activity by capillary zone electrophoresis combined with laser-induced fluorescence.

Skin aging is a progressive process determining the ultimate skin appearance. Human neutrophil elastase (HNE) has been shown to play an important role in the degradation of the extracellular matrix. In order to assay HNE kinetics, a novel online capillary zone electrophoresis (CZE) assay has been developed in this study for the determination of the maximum velocity (Vmax) and of the Michaelis-Menten constant (Km) of HNE regarding several potential substrates. These assays are based on short-end injection to shorten analysis time, on transverse diffusion of laminar flow profiles (TDLFP) for in-capillary reactant mixing, and on UV or laser-induced fluorescence (LIF) detection. Kinetic constants for a referenced peptidic substrate were determined using not only online assays but also offline (pre-capillary) mode. The results obtained were cross compared and compared to the literature in order to validate the developed assays. The hydrolysis of three new potential fluorogenic substrates by HNE was also monitored. Two new peptidic substrates for HNE were identified through this study. Km values of these novel substrates were successfully determined using online CZE assay (Km ∼0.07mM). This value was in the same order of magnitude of that of the referenced substrate despite the presence of the labeling group 5-carboxyfluorescein (5-FAM). HNE activity has never been assessed using online CZE-based assay, neither with UV nor with LIF detection. The developed assay conducted with the new labeled substrates is particularly sensitive (LOQ of few nM), does not require the presence of micelles in the BGE (which is the case for the reference substrate) and only necessitates few nanoliters of reactants making it particularly adapted for screening studies.

Advances in capillary electrophoresis for miniaturizing assays on kinase enzymes for drug discovery.

CE has become a frequently used tool for miniaturizing enzyme assays due particularly to its well-recognized low sample consumption. CE-based enzyme assays cover all aspects of kinetic analysis including the evaluation of enzyme activity, substrate and modulator characterization, and identification. These assays are performed to conduct high-quality primary (hit finding) and secondary (confirmation by determining the half maximal inhibitory concentration, IC50 ) screening. Nowadays, the kinase family is among the most actively studied pharmaceutical targets in oncology, and in neurodegenerative and inflammation diseases. In this article, we review the fundamentals of the different approaches that may be employed for assaying kinase kinetics. Their advantages and limitations will also be discussed by covering the literature of CE-based assays for purified kinases as well as for kinases in living cells. The last section will be devoted to perspectives by showing some applications of multiplexed and of microchip devices for high-throughput screening kinase assays.

New methods based on capillary electrophoresis for in vitro evaluation of protein tau phosphorylation by glycogen synthase kinase 3-ß.

The hyperphosphorylation of tau protein is associated with the development of the neuronal pathology of Alzheimer's disease. As most conventional methods study only particular phosphorylation sites of tau, it is necessary to develop a simple and reliable assay to determine the phosphorylation of tau at multiple sites. Capillary electrophoresis (CE)-based enzymatic assays are not yet used to monitor tau phosphorylation. The present work aims to develop CE-based assays to evaluate tau phosphorylation by the glycogen synthase kinase 3-ß (GSK3ß). A novel pre-capillary CE assay was first developed. An in-capillary CE-based enzymatic assay was also used since this approach is known to be time- and cost- effective. The enzymatic reaction was monitored by quantifying the product adenosine 5'- diphosphate (ADP). The influence of two classes of glycosaminoglycan (GAG), namely heparin and heparan sulfate, on the phosphorylation reaction was also assessed. Results obtained by both CE approaches were comparable and in excellent agreement with those reported in the literature using conventional radiometric and immunoblotting methods. In fact, CE results confirmed the inductory effect of the sulfated sugars heparin and heparan sulfate on tau hyperphosphorylation, probably because of the exposition of new sites phosphorylatable by GSK3ß. This study shows that simple (no-labeling), rapid (less than 30 min per assay), and eco-friendly (no-radioactivity) CE-based kinase assays can give insight into the abnormal phosphorylation of tau. They can be extended to screen different modulators of tau phosphorylation to highlight their function and to develop effective drugs for neurodegenerative disease treatments.