{"id":38958,"date":"2020-01-14T14:55:12","date_gmt":"2020-01-14T12:55:12","guid":{"rendered":"https:\/\/www.iemn.fr\/?p=38958"},"modified":"2020-01-14T14:59:31","modified_gmt":"2020-01-14T12:59:31","slug":"38958","status":"publish","type":"post","link":"https:\/\/www.iemn.fr\/en\/articles-temporaires\/38958.html","title":{"rendered":""},"content":{"rendered":"<div id='layer_slider_1'  class='avia-layerslider main_color avia-shadow  avia-builder-el-0  el_before_av_heading  avia-builder-el-first  container_wrap sidebar_right'  style='height: 261px;'  ><div id=\"layerslider_58_150emiw7bpb1x\" data-ls-slug=\"homepageslider\" class=\"ls-wp-container fitvidsignore ls-selectable\" style=\"width:1140px;height:260px;margin:0 auto;margin-bottom: 0px;\"><div class=\"ls-slide\" data-ls=\"duration:6000;transition2d:5;\"><img loading=\"lazy\" decoding=\"async\" width=\"2600\" height=\"270\" src=\"https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1.jpg\" class=\"ls-bg\" alt=\"\" srcset=\"https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1.jpg 2600w, https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1-300x31.jpg 300w, https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1-768x80.jpg 768w, https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1-1030x107.jpg 1030w, https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1-1500x156.jpg 1500w, https:\/\/www.iemn.fr\/wp-content\/uploads\/2019\/01\/sliders_news1-705x73.jpg 705w\" sizes=\"auto, (max-width: 2600px) 100vw, 2600px\" \/><ls-layer style=\"font-size:14px;text-align:left;font-style:normal;text-decoration:none;text-transform:none;font-weight:700;letter-spacing:0px;border-style:solid;border-color:#000;background-position:0% 0%;background-repeat:no-repeat;width:180px;height:30px;left:0px;top:231px;line-height:32px;color:#ffffff;border-radius:6px 6px 6px 6px;padding-left:50px;background-color:rgba(0, 0, 0, 0.57);\" class=\"ls-l ls-ib-icon ls-text-layer\" data-ls=\"minfontsize:0;minmobilefontsize:0;\"><i class=\"fa fa-quote-right\" style=\"color:#ffffff;margin-right:0.8em;font-size:1em;transform:translateY( -0.125em );\"><\/i>ACTUALITES<\/ls-layer><\/div><\/div><\/div><div id='after_layer_slider_1'  class='main_color av_default_container_wrap container_wrap sidebar_right'  ><div class='container av-section-cont-open' ><div class='template-page content  av-content-small alpha units'><div class='post-entry post-entry-type-page post-entry-38958'><div class='entry-content-wrapper clearfix'>\n\n<style type=\"text\/css\" data-created_by=\"avia_inline_auto\" id=\"style-css-av-k5dvbfuq-07e27ae7392bb879b73d401a2d5f3674\">\n#top .av-special-heading.av-k5dvbfuq-07e27ae7392bb879b73d401a2d5f3674{\nmargin:0 0 10px 0;\npadding-bottom:4px;\n}\nbody .av-special-heading.av-k5dvbfuq-07e27ae7392bb879b73d401a2d5f3674 .av-special-heading-tag .heading-char{\nfont-size:25px;\n}\n.av-special-heading.av-k5dvbfuq-07e27ae7392bb879b73d401a2d5f3674 .av-subheading{\nfont-size:15px;\n}\n<\/style>\n<div  class='av-special-heading av-k5dvbfuq-07e27ae7392bb879b73d401a2d5f3674 av-special-heading-h2  avia-builder-el-1  el_after_av_layerslider  el_before_av_hr  avia-builder-el-first'><h2 class='av-special-heading-tag'  itemprop=\"headline\"  >THESE : Ioana Silvia HOSU &#8211; Ultrasensitive detection of ricin-like proteins by innovative graphene-based sensors, using mass spectrometry<\/h2><div class=\"special-heading-border\"><div class=\"special-heading-inner-border\"><\/div><\/div><\/div>\n\n<style type=\"text\/css\" data-created_by=\"avia_inline_auto\" id=\"style-css-av-18u73nj-dad6a947580930e400fc42ba200e80f1\">\n#top .hr.av-18u73nj-dad6a947580930e400fc42ba200e80f1{\nmargin-top:5px;\nmargin-bottom:5px;\n}\n.hr.av-18u73nj-dad6a947580930e400fc42ba200e80f1 .hr-inner{\nwidth:100%;\n}\n<\/style>\n<div  class='hr av-18u73nj-dad6a947580930e400fc42ba200e80f1 hr-custom  avia-builder-el-2  el_after_av_heading  el_before_av_textblock  hr-left hr-icon-no'><span class='hr-inner inner-border-av-border-thin'><span class=\"hr-inner-style\"><\/span><\/span><\/div>\n<section  class='av_textblock_section av-jriy64i8-2f4600354c0449b610997916bbd9b6bc'   itemscope=\"itemscope\" itemtype=\"https:\/\/schema.org\/BlogPosting\" itemprop=\"blogPost\" ><div class='avia_textblock'  itemprop=\"text\" >\n<style type=\"text\/css\" data-created_by=\"avia_inline_auto\" id=\"style-css-av-13ewzjw-68e036126b913e5028f77311dc66b825\">\n.av_font_icon.av-13ewzjw-68e036126b913e5028f77311dc66b825{\ncolor:#bfbfbf;\nborder-color:#bfbfbf;\n}\n.av_font_icon.av-13ewzjw-68e036126b913e5028f77311dc66b825 .av-icon-char{\nfont-size:60px;\nline-height:60px;\n}\n<\/style>\n<span  class='av_font_icon av-13ewzjw-68e036126b913e5028f77311dc66b825 avia_animate_when_visible av-icon-style- avia-icon-pos-left avia-icon-animate'><span class='av-icon-char' aria-hidden='true' data-av_icon='\ue8c9' data-av_iconfont='entypo-fontello' ><\/span><\/span>\n<p><strong>Ioana Silvia HOSU<\/strong><\/p>\n<p>Soutenance : 11 f\u00e9vrier 2020<strong><br \/>\n<\/strong>IEMN Amphitheatre - Central Laboratory - Villeneuve d'Ascq<\/p>\n<\/div><\/section>\n<section  class='av_textblock_section av-jtefqx33-628129dba2299b2ecd65ebfc92eac29d'   itemscope=\"itemscope\" itemtype=\"https:\/\/schema.org\/BlogPosting\" itemprop=\"blogPost\" ><div class='avia_textblock'  itemprop=\"text\" ><div  class='hr av-kjh3zw-4dff888f744b728a1aca9b3a0971493a hr-default  avia-builder-el-6  avia-builder-el-no-sibling'><span class='hr-inner'><span class=\"hr-inner-style\"><\/span><\/span><\/div>\n<h5><strong><span style=\"color: #800000;\">Jury :<\/span><\/strong><\/h5>\n<div class=\"page\" title=\"Page 1\">\n<div class=\"layoutArea\">\n<div class=\"column\">\n<ul>\n<li>Tuami Lasri, Professeur Universite\u0301 de Lille, IEMN, Examinateur<\/li>\n<li>Celine Elie-Caille, Mai\u0302tre de confe\u0301rences, Universite\u0301 de Franche-Comte\u0301, FEMTO-ST, Rapporteur<\/li>\n<li>Yann Chevolot, Directeur de recherche CNRS, Ecole centrale de Lyon, INL, Rapporteur<\/li>\n<li>Rabah Boukherroub, Directeur de recherche CNRS, Universite\u0301 de Lille, IEMN, Examinateur<\/li>\n<li>Claudia Muracciole Bich, Mai\u0302tre de confe\u0301rences, Universite\u0301 de Montpellier, IBMM, Examinateur<\/li>\n<li>Yannick Coffinier, Charg\u00e9 de recherche CNRS, Universite\u0301 de Lille, IEMN, Directeur de th\u00e8se<\/li>\n<li>Didier Hilaire, DGA, Paris, Invit\u00e9<\/li>\n<\/ul>\n<\/div>\n<\/div>\n<\/div>\n<h5>Summary:<\/h5>\n<div class=\"page\" title=\"Page 5\">\n<div class=\"layoutArea\">\n<div class=\"column\">\n<p>Les attaques bioterroristes sont devenues plus fre\u0301quentes ces dernie\u0300res anne\u0301es et le large e\u0301ventail d\u2019agents bioterroristes en fait un proble\u0300me important a\u0300 re\u0301soudre. La ricine appartient a\u0300 la famille des prote\u0301ines inactivant les ribosomes (RIP). Les RIP sont des toxines biologiques, solubles dans l\u2019eau, qui peuvent e\u0302tre facilement extraites de plantes (ricine de graines de Ricinus communis et abrine de graines d\u2019Abrus precatorius) ou de bacte\u0301ries (toxine de Shiga). La ricine est compose\u0301e de deux chai\u0302nes: la chai\u0302ne A de la ricine, une N- glycosidase induisant la toxicite\u0301 par e\u0301limination de l\u2019ade\u0301nine (action de de\u0301purination) de l\u2019ARNr 28S des sous-unite\u0301s ribosomales 60S, inhibant la synthe\u0300se prote\u0301ique, et la chai\u0302ne B, une lectine qui se lie aux fragments de sucre spe\u0301cifiques sur la membrane extracellulaire, assurant l&rsquo;absorption de la toxine. Comme ils inhibent la synthe\u0300se des prote\u0301ines, en fonction de la voie d&rsquo;absorption (orale, par inhalation, par voie intraveineuse) et de la dose rec\u0327ue, la mort peut survenir. En l&rsquo;absence de contre-mesures efficaces, les me\u0301thodes de de\u0301tection de ces toxines doivent e\u0302tre rapides, fiables, se\u0301lectives et sans aucune ambigui\u0308te\u0301. Les me\u0301thodes actuelles qui sont principalement base\u0301es sur des me\u0301thodes comme le SERS, l\u2019ELISA, la Colorime\u0301trie et la SPR ne re\u0301pondent pas a\u0300 toutes ces exigences.<\/p>\n<p>Me\u0302me si la spectrome\u0301trie de masse a e\u0301te\u0301 utilise\u0301e pour la de\u0301tection de la ricine, elle ne peut pas e\u0302tre re\u0301alise\u0301e sans une longue et fastidieuse pre\u0301paration d&rsquo;e\u0301chantillon, c&rsquo;est-a\u0300-dire une digestion enzymatique, une extraction \/purification et une identification de prote\u0301ines sur la base d&rsquo;une analyse peptidique suivie d&rsquo;une comparaison avec des bases de donne\u0301es existantes. Dans ce travail, nous avons montre\u0301 comment les mate\u0301riaux a\u0300 base de carbone (nanomurs de carbone) pourraient e\u0302tre applique\u0301s comme mate\u0301riaux nanostructure\u0301s pour la de\u0301tection de la ricine par de\u0301sorption\/ionisation laser de surface pour la de\u0301tection par spectrome\u0301trie de masse (SALDI-MS). Tout d&rsquo;abord, l&rsquo;ade\u0301quation des nanoparticules de carbone en tant que bonne surface SALDI a e\u0301te\u0301 initialement e\u0301tudie\u0301e pour des biomole\u0301cules plus petites (saccharides, peptides jusqu\u2019a\u0300 5800 m \/ z, glucides, lipides et glucose, qui a e\u0301galement e\u0301te\u0301 quantifie\u0301 a\u0300 l\u2019aide de SALDI-MS).<\/p>\n<p>En ce qui concerne les prote\u0301ines, la litte\u0301rature a montre\u0301 qu&rsquo;elles sont difficiles a\u0300 ioniser et a\u0300 de\u0301tecter avec la me\u0301thode SALD-MS, en raison de leur grand poids mole\u0301culaire. La capacite\u0301 des CNWs a\u0300 de\u0301sorber et a\u0300 ioniser les prote\u0301ines a ne\u0301cessite\u0301 de nombreuses e\u0301tapes d\u2019optimisation (re\u0301alise\u0301e pour la de\u0301tection de prote\u0301ines en jouant avec la nature et la concentration de sel, le temps d\u2019incubation, les caracte\u0301ristiques physico-chimiques des nanomurs (comme la hauteur, le dopage au bore ou la morphologie). Pour ce faire, le cytochrome C a e\u0301te\u0301 utilise\u0301 comme prote\u0301ine mode\u0300le. Enfin, des nanomurs de carbone aligne\u0301es verticalement ont ensuite e\u0301te\u0301 modifie\u0301es a\u0300 l&rsquo;aide de sucres a\u0300 lectine spe\u0301cifiques (galactosamine), pour la de\u0301tection spe\u0301cifique de la chai\u0302ne B de la ricine dans des e\u0301chantillons re\u0301els, tels que des boissons sans alcool et du se\u0301rum sanguin. Nous avons obtenu une limite de de\u0301tection (80 ng\/0.5 \u03bcL) soit trois fois infe\u0301rieure a\u0300 la dose le\u0301tale me\u0301diane la plus faible (DL50 = 10 \u03bcg\/kg). Cette de\u0301tection peut e\u0302tre re\u0301alise\u0301e dans les 10 min.<\/p>\n<\/div>\n<\/div>\n<\/div>\n<div class=\"page\" title=\"Page 6\">\n<div class=\"layoutArea\">\n<div class=\"column\">\n<p>Dans la dernie\u0300re partie, des re\u0301sultats pre\u0301liminaires concernant la mise au point d&rsquo;outils analytiques bimodaux seront pre\u0301sente\u0301s. Il s&rsquo;agit de combinaisons telles que: SPR (re\u0301sonance plasmonique de surface)\/SALDI-MS, SERS (Spectroscopie Raman Exalte\u0301e de Surface)\/SALDI-MS et EC(E\u0301lectrochimie)\/SALDI-MS. Une attention particulie\u0300re a e\u0301te\u0301 porte\u0301e sur la SPR \/ SALDI-MS car elle permet d\u2019obtenir des interactions quantitatives et mole\u0301culaires en temps re\u0301el (SPR) et une identification structurelle des analytes (MS). Diffe\u0301rentes me\u0301thodes de de\u0301po\u0302t de mate\u0301riaux de type graphe\u0300ne ont e\u0301te\u0301 e\u0301tudie\u0301es (me\u0301thode de surfactant a\u0300 bulle d&rsquo;oxyde de graphe\u0300ne, transfert par voie humide de graphe\u0300ne CVD, de\u0301po\u0302t e\u0301lectrophore\u0301tique de graphe\u0300ne, couche par couche en utilisant un polycation et un oxyde de graphe\u0300ne, coule\u0301e en goutte a\u0300 goutte de graphe\u0300ne et d&rsquo;oxyde de graphe\u0300ne re\u0301duit). Le cytochrome C (en tant que prote\u0301ine mode\u0300le) a e\u0301te\u0301 de\u0301tecte\u0301 en utilisant les trois premie\u0300res me\u0301thodes et seuls les peptides ont e\u0301te\u0301 de\u0301tecte\u0301s pour les deux dernie\u0300res me\u0301thodes, l&rsquo;ordre de mention e\u0301tant en corre\u0301lation avec la diminution de l&rsquo;efficacite\u0301 de SALDI-MS vis-a\u0300-vis des prote\u0301ines.<\/p>\n<p>Cette the\u0300se de\u0301crit pour la premie\u0300re fois le de\u0301veloppement d&rsquo;un capteur de type SALDI- MS, capable de de\u0301tecter la ricine a\u0300 une dose infe\u0301rieure a\u0300 la dose mortelle chez l&rsquo;homme et d&rsquo;apporter ainsi une contribution importante a\u0300 la lutte contre d&rsquo;e\u0301ventuelles attaques terroristes. L&rsquo;e\u0301tude syste\u0301matique de diffe\u0301rents parame\u0300tres qui influencent ce processus LDI-MS est e\u0301galement pre\u0301sente\u0301. Les techniques bimodales comme la SPR-MS, pre\u0301sentent des alternatives inte\u0301ressantes permettant de cre\u0301er des outils analytiques plus puissants.<\/p>\n<\/div>\n<\/div>\n<\/div>\n<h5>Abstract:<\/h5>\n<div class=\"page\" title=\"Page 3\">\n<div class=\"layoutArea\">\n<div class=\"column\">\n<p>Bio-terroristic attacks have become more frequent in the past years and the wide range of bio-terroristic agents makes this an important issue to overcome. Ricin is part of the ribosome-inactivating proteins (RIP). RIPs are vegetable toxins, water soluble, which can be easily extracted from plants (ricin from castor beams, abrin from rosary pea) or from bacteria (Shiga toxin). These proteins are composed of two chains: ricin A chain, a glycosidase that insures the toxicity by removal of adenine (depurination) from the RNAr 28S from the 60S ribosomal subunits, followed by the inhibition of protein synthesis, and ricin B chain, a lectin that binds to specific sugar moieties on the surface of the cells, assuring transportation the cell uptake. As they inhibit protein synthesis, depending of the administration take-up (oral, inhalation, intravenously) and the dose received, cell death also occurs. In the absence of efficient counter measurements, detection methods of these toxins have to be fast, reliable, selective and suitable, especially pre-assimilation analysis. The current methods (based on SERS, ELISA, Colorimetric, SPR and MS) do not overcome all these requirements.<\/p>\n<p>Even though mass spectrometry was used for ricin detection, it cannot be performed without long and tedious sample preparation, meaning through enzymatic digestion, extraction\/purification and identification of the proteins based on the peptide analysis, followed by comparison with an existent database (mass fingerprint). In this work, we describe how carbon-based materials (carbon nanowalls and others) can be used as nanostructured materials for specific ricin-like proteins sensors, using surface assisted laser\/desorption ionization mass spectrometry (SALDI-MS) and other techniques. The suitability of the carbon nanowalls (CNWS) was proven initially for other smaller bio- molecules (saccharides, peptides up to 5800 m\/z, carbohydrates, lipids and glucose, which was also quantified using SALDI-MS).<\/p>\n<p>When it comes to proteins, they are hard to ionize and detect using SALD-MS, due in part to their big molecular weight. The ability of CNWs to desorb and ionize proteins required a lot of optimization steps of the SALDI-MS method (salt nature, concentration, and pH, incubation time, physicochemical characteristics of the nanowalls, such as height, boron doping and morphology). A systematic optimization was done using a model protein, the cytochrome C). From this, we were able, for the first time, to detect Ricin B chain without the use of organic matrix. To go further in improving Ricin detection performances, carbon nanowalls were then covalently modified using specific lectin sugars (galactosamine) and the ability to detect Ricin B chain in real samples such as soft drinks and blood serum was demonstrated within10 minutes. We obtained a limit of detection (80 ng\/0.5 \u03bcL) that is 3 times lower than the lowest median lethal dose (LD50 = 10 \u03bcg\/kg)<\/p>\n<\/div>\n<\/div>\n<\/div>\n<div class=\"page\" title=\"Page 4\">\n<div class=\"layoutArea\">\n<div class=\"column\">\n<p>Multifunctional surfaces are described as perspectives for more powerful bimodal analytical tools, by combining two techniques, such as: SPR(Surface Plasmon Resonance)\/SALDI-MS, SERS(Surface Enhanced Raman Spectroscopy)\/SALDI-MS and EC(Electrochemistry)\/SALDI-MS. Special attention was focused on SPR\/SALDI-MS as it can achieve both quantitative and molecular interactions in real-time (SPR) and precise identification of the analytes (MS). Different depositions methods of graphene-like materials were studied to ensure a good surface coverage of the substrate : 1)bubble surfactant method of graphene oxide, 2) wet transfer of CVD pristine graphene, 3) electrophoretic deposition of graphene, 4) layer by layer using a polycation and graphene oxide, 5) drop casting of both graphene and reduced graphene oxide). Cytochrome C (as model protein) was detected using the first three methods and only peptides were detected for the last two methods, as the mentioning order is in correlation with decreasing SALDI-MS efficiency towards proteins.<\/p>\n<p>In this thesis, we described the first world wide ricin-like proteins SALDI-MS sensor, which is able to detect below the lethal dose in humans and bring an important contribution to the fight against eventual terroristic attacks. The systematic study of different parameters that influence this LDI-MS process is also presented. The dual surfaces studied, in particular the SPR\/MS bimodal techniques, presented reliable consistency for further approaches in creating more powerful analytical tools.<\/p>\n<\/div>\n<\/div>\n<\/div>\n<\/div><\/section>","protected":false},"excerpt":{"rendered":"","protected":false},"author":2,"featured_media":0,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[36],"tags":[],"class_list":["post-38958","post","type-post","status-publish","format-standard","hentry","category-articles-temporaires"],"_links":{"self":[{"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/posts\/38958","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/users\/2"}],"replies":[{"embeddable":true,"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/comments?post=38958"}],"version-history":[{"count":0,"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/posts\/38958\/revisions"}],"wp:attachment":[{"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/media?parent=38958"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/categories?post=38958"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.iemn.fr\/en\/wp-json\/wp\/v2\/tags?post=38958"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}