{"id":1010,"date":"2017-02-27T21:26:38","date_gmt":"2017-02-27T21:26:38","guid":{"rendered":"https:\/\/www.progeriaresearch.org\/?page_id=1010"},"modified":"2026-03-17T10:50:58","modified_gmt":"2026-03-17T14:50:58","slug":"induced-pluripotent-stem-cells","status":"publish","type":"page","link":"https:\/\/www.progeriaresearch.org\/fr\/induced-pluripotent-stem-cells\/","title":{"rendered":"Cellules souches pluripotentes induites"},"content":{"rendered":"

[et_pb_section fb_built=\u201d1\u2033 fullwidth=\u201don\u201d disabled_on=\u201doff|off|off\u201d _builder_version=\u201d4.16\u2033 border_width_bottom=\u201d55px\u201d border_color_bottom=\u201d#29327a\u201d locked=\u201doff\u201d global_colors_info=\u201d{}\u201d][et_pb_fullwidth_header _builder_version=\u201d4.16\u2033 title_font=\u201d||||||||\u201d title_font_size=\u201d55\u2033 background_color=\u201d#29327a\u201d background_image=\u201dhttps:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2020\/12\/Jes.jpg\u201d background_position=\u201dcenter_left\u201d custom_padding=\u201d9vw|0px|9vw||true|\u201d custom_padding_tablet=\u201d\u201d custom_padding_phone=\u201d|56px||\u201d custom_padding_last_edited=\u201dsur|bureau\u201d title_font_size_tablet=\u201d45px\u201d title_font_size_phone=\u201d40px\u201d title_font_size_last_edited=\u201dsur|t\u00e9l\u00e9phone\u201d z_index_tablet=\u201d500\u2033 custom_css_main_element=\u201dbackground-position: center 18% !important;\u201d global_colors_info=\u201d{}\u201d]<\/p>\n

Pluripotent induit<\/h1>\n

Cellules souches<\/h1>\n

 <\/p>\n

[\/et_pb_fullwidth_header][\/et_pb_section][et_pb_section fb_built=”1″ use_custom_gutter=”on” gutter_width=”1″ specialty=”on” padding_left_1=”35px” padding_left_2=”35px” padding_2_tablet=”|||0px” padding_2_phone=”” padding_2_last_edited=”on|desktop” module_class_1=”sidebar-secondary-nav” module_class=”handprint-bg” _builder_version=”4.16″ background_image=”https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/04\/blue-handprint-only.png” parallax=”on” parallax_method=”off” inner_width=”100%” inner_max_width=”100%” custom_padding=”0|0px|54px|0px|false|false” z_index_tablet=”500″ border_width_top=”10px” border_color_top=”#8fd2ed” use_custom_width=”on” width_unit=”off” custom_width_percent=”100%” global_colors_info=”{}”][et_pb_column type=”1_4″ _builder_version=”4.16″ custom_padding=”|||” global_colors_info=”{}” custom_padding__hover=”|||”][et_pb_sidebar area=”et_pb_widget_area_14″ disabled_on=”on|on|off” module_class=”subpage-sidebars” _builder_version=”4.16″ animation_style=”fade” z_index_tablet=”500″ border_width_right=”5px” locked=”off” global_colors_info=”{}”][\/et_pb_sidebar][\/et_pb_column][et_pb_column type=”3_4″ specialty_columns=”3″ _builder_version=”4.16″ custom_padding=”|||” global_colors_info=”{}” custom_padding__hover=”|||”][et_pb_row_inner custom_padding_last_edited=”on|phone” _builder_version=”4.27.5″ custom_padding=”|35px|0|0px|false|false” custom_padding_tablet=”|35px||35px||true” custom_padding_phone=”” animation_direction=”top” global_colors_info=”{}”][et_pb_column_inner saved_specialty_column_type=”3_4″ _builder_version=”4.16″ custom_padding=”|||” global_colors_info=”{}” custom_padding__hover=”|||”][et_pb_text _builder_version=”4.27.5″ background_size=”initial” background_position=”top_left” background_repeat=”repeat” custom_padding=”||0px||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

La Banque de cellules et de tissus de la Fondation de recherche sur la prog\u00e9ria<\/strong> Cellules souches pluripotentes induites humaines (iPSC)<\/h4>\n
    \n
  1. Cellules iPSC de la prog\u00e9ria <\/strong>Informations g\u00e9n\u00e9rales pour les non-scientifiques<\/strong><\/a>\u00a0<\/strong><\/li>\n
  2. Objectif de <\/strong>G\u00e9n\u00e9ration et distribution de cellules souches pluripotentes induites par la Progeria Research Foundation\u00a0<\/strong>\u00a0<\/a><\/li>\n
  3. G\u00e9n\u00e9ration de cellules souches pluripotentes induites par le syndrome de Hutchinson-Gilford-Progeria (iPSC)<\/strong>\u00a0<\/a><\/li>\n
  4. Contr\u00f4le qualit\u00e9 : validation et caract\u00e9risation<\/strong>\u00a0<\/a><\/li>\n
  5. Mat\u00e9riau de d\u00e9part d'origine \u00e0 partir duquel les iPSC ont \u00e9t\u00e9 d\u00e9riv\u00e9es<\/strong><\/a><\/li>\n
  6. Rejoignez notre liste de diffusion pour les futures mises \u00e0 jour sur les iPSC et les nouvelles lign\u00e9es cellulaires<\/strong>\u00a0\u00a0<\/strong><\/a><\/li>\n
  7. Des questions ? Contactez-nous.<\/strong><\/a><\/li>\n
  8. Commande de lignes iPSC<\/strong><\/a>\u00a0<\/strong><\/li>\n
  9. Pr\u00e9paration des milieux de culture HGPS et iPSC de contr\u00f4le<\/strong>\u00a0<\/strong><\/a><\/li>\n
  10. Pr\u00e9paration des plaques de Matrigel\u00a0<\/strong><\/a><\/li>\n
  11. D\u00e9cong\u00e9lation des cellules hESC et iPSC (par cryo-flacon)<\/strong><\/a><\/li>\n
  12. R\u00e9colte et entretien des cellules souches embryonnaires humaines (hESC) et cellules souches embryonnaires pluripotentes induites (iPSC)<\/strong>\u00a0<\/strong><\/a><\/li>\n
  13. Passage <\/strong><\/a>hESC\/iPSC\u00a0<\/a><\/strong><\/li>\n
  14. Cong\u00e9lation des cellules souches embryonnaires humaines (hESC) et cellules souches embryonnaires in vitro (iPSC)<\/a><\/strong><\/li>\n<\/ol>\n

    [\/et_pb_text][\/et_pb_column_inner][\/et_pb_row_inner][et_pb_row_inner custom_padding_last_edited=”on|phone” _builder_version=”4.27.5″ custom_padding=”42px|35px|0|0px|false|false” custom_padding_tablet=”|35px||35px||true” custom_padding_phone=”” animation_direction=”top” border_width_top=”10px” border_color_top=”#29327a” global_colors_info=”{}”][et_pb_column_inner saved_specialty_column_type=”3_4″ _builder_version=”4.16″ custom_padding=”|||” global_colors_info=”{}” custom_padding__hover=”|||”][et_pb_text admin_label=”iPSC” module_id=”iPSC” _builder_version=”4.27.5″ custom_padding=”||0px||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

    1. Informations g\u00e9n\u00e9rales sur les iPSC pour les non-scientifiques<\/strong><\/h4>\n

    Les cellules souches sont des cellules \u00ab immatures \u00bb qui n\u2019ont pas encore pris l\u2019engagement de devenir un type cellulaire particulier. Elles sont mall\u00e9ables car elles ont le potentiel de se d\u00e9velopper en de nombreux types diff\u00e9rents de cellules matures dans le corps, comme les cellules qui composent le c\u0153ur ou les vaisseaux sanguins, ainsi que d\u2019autres tissus et organes. En 2007, des chercheurs ont d\u00e9couvert une strat\u00e9gie pour cr\u00e9er des cellules souches en laboratoire en reprogrammant des cellules adultes matures que nous cultivons g\u00e9n\u00e9ralement \u00e0 des fins de recherche.1, 2<\/sup><\/strong>\u00a0Ces cellules souches artificiellement cr\u00e9\u00e9es sont appel\u00e9es cellules souches pluripotentes induites (\u00ab iPSC \u00bb). Pour le domaine de la prog\u00e9ria, il s\u2019agit d\u2019une avanc\u00e9e majeure. Pour la premi\u00e8re fois, les scientifiques peuvent d\u00e9sormais fabriquer des cellules souches de la prog\u00e9ria et se demander comment les cellules souches fonctionnent et se d\u00e9veloppent dans la prog\u00e9ria. Jusqu\u2019\u00e0 pr\u00e9sent, il n\u2019existait aucune source de cellules souches humaines de la prog\u00e9ria, et il y avait donc un manque d\u2019informations sur le fonctionnement des cellules souches de la prog\u00e9ria par rapport aux cellules souches de personnes non atteintes de la prog\u00e9ria. En outre, les scientifiques peuvent reprogrammer les cellules souches de la prog\u00e9ria pour cr\u00e9er, pour la premi\u00e8re fois, des cellules vasculaires, cardiaques et d\u2019autres types de cellules matures de la prog\u00e9ria. Jusqu\u2019\u00e0 pr\u00e9sent, il n\u2019existait aucune source de cellules cardiaques ou vasculaires humaines de la prog\u00e9ria.\u00a0\u00a0Nous pouvons maintenant demander la cl\u00e9\u00a0<\/strong>Des questions se posent sur la maladie cardiaque qui entra\u00eene une mort pr\u00e9matur\u00e9e chez les personnes atteintes de prog\u00e9ria, par crise cardiaque ou accident vasculaire c\u00e9r\u00e9bral. Nous pouvons comparer ces d\u00e9couvertes avec les maladies cardiaques et le vieillissement dans la population g\u00e9n\u00e9rale et en savoir plus sur ce qui influence le vieillissement chez nous tous. Plusieurs excellentes \u00e9tudes ont d\u00e9j\u00e0 \u00e9t\u00e9 publi\u00e9es sur les cellules souches de la prog\u00e9ria.3-5<\/sup> \u00a0Notre objectif \u00e0 la Progeria Research Foundation est de faciliter de nombreuses autres d\u00e9couvertes gr\u00e2ce \u00e0 cet outil pr\u00e9cieux. Pour une introduction aux cellules souches, veuillez consulter ce site Web du gouvernement am\u00e9ricain :\u00a0https:\/\/stemcells.nih.gov\/info\/basics.htm<\/a><\/p>\n

     <\/p>\n

    \n
      \n
    1. \n
        \n
      1. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S. Induction de cellules souches pluripotentes \u00e0 partir de fibroblastes humains adultes par des facteurs d\u00e9finis.\u00a0Cellule.\u00a0<\/em>2007;131:861-872.<\/li>\n
      2. Yu J, Vodyanik MA, Smuga-Otto K, Antosiewicz-Bourget J, Frane JL, Tian S, Nie J, Jonsdottir GA, Ruotti V, Stewart R, Slukvin, II, Thomson JA. Lign\u00e9es de cellules souches pluripotentes induites d\u00e9riv\u00e9es de cellules somatiques humaines.\u00a0Science.\u00a0<\/em>2007;318:1917-1920.<\/li>\n
      3. Liu GH, Barkho BZ, Ruiz S, Diep D, Qu J, Yang SL, Panopoulos AD, Suzuki K, Kurian L, Walsh C, Thompson J, Boue S, Fung HL, Sancho-Martinez I, Zhang K, Yates J, 3rd, Izpisua Belmonte JC. R\u00e9capitulation du vieillissement pr\u00e9matur\u00e9 avec les iPSC du syndrome de prog\u00e9ria de Hutchinson-Gilford.\u00a0Nature.\u00a0<\/em>2011;472:221-225.<\/li>\n
      4. Misteli T. iPSCs d\u00e9riv\u00e9es de HGPS pour les \u00e2ges.\u00a0Cellule souche.\u00a0<\/em>2011;8:4-6.<\/li>\n<\/ol>\n<\/li>\n<\/ol>\n<\/div>\n

        [\/et_pb_text][et_pb_text admin_label=”purpose” module_id=”purpose” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

        2. Objectif de la g\u00e9n\u00e9ration et de la distribution de cellules souches pluripotentes induites (iPSC) par la Progeria Research Foundation<\/strong><\/h4>\n

        La mission de la Fondation de recherche sur la Prog\u00e9ria est de d\u00e9couvrir des traitements et des rem\u00e8des contre le syndrome de Hutchinson-Gilford de la Prog\u00e9ria et les troubles li\u00e9s au vieillissement. En 2009, la PRF a conclu une collaboration avec une \u00e9quipe d'experts scientifiques de l'Universit\u00e9 de Toronto, au Canada, sous la direction du Dr William Stanford, afin de g\u00e9n\u00e9rer des cellules souches embryonnaires pluripotentes induites (iPSC) de la Prog\u00e9ria de haute qualit\u00e9. Le Dr Stanford est titulaire de la chaire de recherche du Canada en biologie int\u00e9grative des cellules souches. Depuis 2011, la PRF continue de collaborer avec le Dr Stanford \u00e0 l'Universit\u00e9 d'Ottawa, au Canada, o\u00f9 il est professeur de m\u00e9decine cellulaire et mol\u00e9culaire \u00e0 la Facult\u00e9 de m\u00e9decine et chercheur principal au Centre Sprott de recherche sur les cellules souches de l'Institut de recherche de l'h\u00f4pital d'Ottawa.<\/p>\n

        Notre objectif est de fournir cet outil pr\u00e9cieux aux chercheurs du monde entier. Ce nouvel outil de recherche servira \u00e0 g\u00e9n\u00e9rer de nouvelles recherches innovantes sur la prog\u00e9ria, ainsi que sur sa relation avec les maladies cardiaques et le vieillissement.\u00a0\u00a0<\/strong><\/p>\n

        [\/et_pb_text][et_pb_text admin_label=”generation” module_id=”generation” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

        3. G\u00e9n\u00e9ration de cellules souches pluripotentes induites par le syndrome de Hutchinson-Gilford-Progeria (iPSC)<\/strong><\/h4>\n

        Les cellules souches pluripotentes induites (iPSC) ont \u00e9t\u00e9 d\u00e9riv\u00e9es \u00e0 l'aide de la transduction r\u00e9trovirale pseudotyp\u00e9e VSVG de quatre facteurs humains, Oct4, Sox2, Klf4 et c-Myc dans des fibroblastes. Les colonies d'iPSC ont \u00e9t\u00e9 d\u00e9riv\u00e9es sur des fibroblastes embryonnaires de souris (MEF). La proc\u00e9dure utilis\u00e9e \u00e9tait essentiellement celle d\u00e9crite pr\u00e9c\u00e9demment, mais sans l'utilisation du rapporteur EOS (Nature Protocols 4\u00a0: 1828-1844, 2009).<\/p>\n

        [\/et_pb_text][et_pb_text admin_label=”quality” module_id=”quality” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ saved_tabs=”all” global_colors_info=”{}”]<\/p>\n

        4. Contr\u00f4le qualit\u00e9 : validation et caract\u00e9risation<\/strong><\/h4>\n

        Les lignes actuellement disponibles ont subi plusieurs \u00e9tapes de validation (voir PDF t\u00e9l\u00e9chargeables ci-dessous) :<\/p>\n

        \n
          \n
        1. \n
            \n
          1. Test de mycoplasme pour chaque lign\u00e9e : le laboratoire du Dr Stanford a effectu\u00e9 une analyse de mycoplasme par PCR pour chaque lign\u00e9e cellulaire. De plus, apr\u00e8s l'expansion et avant l'exp\u00e9dition des cellules, les lign\u00e9es seront \u00e0 nouveau test\u00e9es pour d\u00e9tecter les mycoplasmes.<\/li>\n
          2. Immunocoloration pour les marqueurs de pluripotence Tra-1-60, Tra-1-81 et SSEA4.<\/li>\n
          3. La coloration \u00e0 la phosphatase alcaline comme indicateur de pluripotence<\/li>\n
          4. Formation du corps embryonnaire et immunocoloration ult\u00e9rieure pour les marqueurs des trois feuillets germinaux. Les marqueurs test\u00e9s \u00e9taient la \u03b2III-tubuline (ectoderme), l'actine des muscles lisses (m\u00e9soderme) et Gata4 ou AFP (endoderme)<\/li>\n
          5. Analyse du caryotype.<\/li>\n
          6. R\u00e9expression de la lamine A dans les cellules diff\u00e9renci\u00e9es<\/li>\n
          7. Essais de t\u00e9ratome<\/li>\n<\/ol>\n<\/li>\n<\/ol>\n

            Validation suppl\u00e9mentaire en cours :<\/strong>
            Certaines lign\u00e9es ont r\u00e9alis\u00e9 des tests de t\u00e9ratome, comme le montrent les donn\u00e9es justificatives. Pour toutes les autres lign\u00e9es, des tests de t\u00e9ratome sont en cours et le statut sera mis \u00e0 jour au fur et \u00e0 mesure que ces tests seront termin\u00e9s.<\/p>\n<\/div>\n

            [\/et_pb_text][et_pb_text admin_label=”original” module_id=”original” _builder_version=”4.27.5″ custom_padding=”||0px|||” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

            5. Mat\u00e9riau de d\u00e9part original \u00e0 partir duquel ces cellules iPS ont \u00e9t\u00e9 d\u00e9riv\u00e9es<\/strong><\/h4>\n

            Les iPSC proviennent de lign\u00e9es cellulaires de fibroblastes non transform\u00e9s de la PRF Cell & Tissue Bank.<\/p>\n

            La m\u00e9thode de transduction utilis\u00e9e pour toutes les lign\u00e9es iPS \u00e9tait le Retrovirus MKOS.<\/p>\n

            <\/p>\n

            [\/et_pb_text][et_pb_text admin_label=”Table” module_id=”original” _builder_version=”4.27.5″ custom_padding=”||0px|||” hover_enabled=”0″ z_index_tablet=”500″ global_colors_info=”{}” sticky_enabled=”0″]<\/p>\n

            \n\n\n\n\t\n\n\t\n\t\n\t\n\t\n\t\n\t\n\t\n\t\n\t
            ID de ligne iPSC<\/b><\/th>Mutation<\/b><\/th>Sexe et \u00e2ge du don<\/b><\/th>Type de cellule d'origine Cliquez ici<\/a>.<\/b><\/th>Donn\u00e9es \u00e0 l'appui<\/b><\/th>\n<\/tr>\n<\/thead>\n
            HGADFN003 iPS 1B<\/td> LMNAExon 11,
            \n1824 C>T<\/td>            		Homme 2 ans 0 mois<\/td> Fibroblastes dermiques
            \nHGADFN003<\/td>            		003 iPS1B<\/a><\/td>\n<\/tr>\n
            HGADFN003 iPS 1C<\/td> Exon 11 de l'ARNm,
            \n1824 C>T<\/td>            		Homme 2 ans 0 mois<\/td> Fibroblastes dermiques
            \nHGADFN003<\/td>            		003 iPS1C<\/a><\/td>\n<\/tr>\n
            HGDFN003
            \niPS 1D<\/td>            		Exon 11 de l'ARNm,
            \n1824 C>T<\/td>            		Homme 2 ans 0 mois<\/td> Fibroblastes dermiques
            \nHGADFN003<\/td>            		003 iPS1D<\/a><\/td>\n<\/tr>\n
            HGADFN167 iPS 1J<\/td> Exon 11 de LMNA, 1824 C>T<\/td>Homme 8 ans 5 mois<\/td>Fibroblastes dermiques HGADFN167<\/td>167 PS 1J<\/a><\/td>\n<\/tr>\n
            HGADFN167 iPS 1Q<\/td> Exon 11 de LMNA, 1824 C>T<\/td>Homme 8 ans 5 mois<\/td>Fibroblastes dermiques HGADFN167<\/td>167 iPS1Q<\/a><\/td>\n<\/tr>\n
            HGMDFN090 iPS 1B<\/td> M\u00e8re de HGADFN167 (non affect\u00e9e)<\/td>Femme 37 ans 10 mois<\/td>Fibroblastes dermiques
            \nHGMDFN090<\/td>            		090 iPS1B<\/a><\/td>\n<\/tr>\n
            HGMDFN090 iPS 1C<\/td> M\u00e8re de HGADFN167 (non affect\u00e9e)<\/td>Femme 37 ans 10 mois<\/td>Fibroblastes dermiques
            \nHGMDFN090<\/td>            		090 iPS1C<\/a><\/td>\n<\/tr>\n
            HGFDFN168 iPS1 D2<\/td>P\u00e8re de HGADFN167 (non affect\u00e9)<\/td>Homme 40 ans
            \n5 mois<\/td>            		Fibroblastes dermiques HGFDFN168<\/td>168 iPS1 D2<\/a><\/td>\n<\/tr>\n
            HGFDFN168 iPS1P<\/td>P\u00e8re de HGADFN167 (non affect\u00e9)<\/td>Homme 40 ans
            \n5 mois<\/td>            		Fibroblastes dermiques
            \nHGFDFN168<\/td>            		168 iPS1P<\/a><\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n<\/h4>\n

            [\/et_pb_text][et_pb_button button_url=”https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2023\/05\/PRF-AVAILABLE-CELL-LINES-5-24-23.pdf” button_text=”PRF Available Cell Lines” button_alignment=”left” admin_label=”Cell lines pdf” _builder_version=”4.27.5″ background_layout=”dark” custom_margin=”||25px||false|false” hover_enabled=”0″ z_index_tablet=”500″ global_colors_info=”{}” sticky_enabled=”0″][\/et_pb_button][et_pb_text admin_label=”join” module_id=”join” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

            6. Rejoignez notre liste de diffusion pour les futures mises \u00e0 jour sur les iPSC et les nouvelles lign\u00e9es cellulaires<\/strong><\/h4>\n

            Nous continuons \u00e0 g\u00e9n\u00e9rer des lign\u00e9es iPSC. Si vous souhaitez recevoir des mises \u00e0 jour p\u00e9riodiques sur les iPSC conserv\u00e9es dans la PRF Cell & Tissue Bank, veuillez vous inscrire \u00e0 notre liste de diffusion en cliquant sur\u00a0ici<\/a><\/strong><\/p>\n

            [\/et_pb_text][et_pb_text admin_label=”questions” module_id=”questions” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

            7. Des questions ?<\/strong><\/h4>\n

            <\/a>Veuillez contacter Leslie Gordon, MD, PhD, directrice m\u00e9dicale, pour toute question ou besoin, au lgordon@progeriaresearch.org<\/a>\u00a0ou 978-535-2594<\/p>\n

            [\/et_pb_text][et_pb_text admin_label=”ordering” module_id=”ordering” _builder_version=”4.27.5″ custom_padding=”||0px||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

            8. Commande de lign\u00e9es cellulaires iPS<\/strong><\/h4>\n

            En 2014, la PRF a instaur\u00e9 une politique de non-modification de son MTA. C'est le r\u00e9sultat de 12 ann\u00e9es d'accords contractuels avec 70 \u00e9quipes de recherche travaillant dans des institutions de 14 pays. La PRF et ses conseillers juridiques ont pris en consid\u00e9ration les probl\u00e8mes survenus au cours de cette p\u00e9riode et ont modifi\u00e9 l'accord en cons\u00e9quence, ce qui nous semble \u00eatre des conditions justes et raisonnables.<\/p>\n

            Pour les institutions du gouvernement f\u00e9d\u00e9ral am\u00e9ricain ou pour toute question, veuillez contacter Wendy Norris \u00e0 : wnorris@brownhealth.org<\/a><\/a>\u00a0ou\u00a0401-274-1122 x 48063<\/a>.<\/p>\n

            \u00c9tape 1 : Remplir une demande et un accord de transfert de mat\u00e9riel
            <\/strong>
            <\/a>            Demande et accord pour les institutions non gouvernementales<\/a><\/p>\n

            Accord de transfert de mat\u00e9riel pour les institutions non gouvernementales<\/a><\/p>\n

            \u00c9tape 2 : Renvoyez la demande compl\u00e9t\u00e9e et l'accord de transfert de mat\u00e9riel \u00e0 Wendy Norris \u00e0 wnorris@brownhealth.org<\/a>Une fois approuv\u00e9e, vous recevrez un e-mail confirmant votre commande et la date d'exp\u00e9dition pr\u00e9vue.<\/b>\u00a0<\/strong><\/a><\/p>\n

            \u00c9tape 3 : <\/strong>Le laboratoire du Dr Stanford distribue actuellement des lign\u00e9es dans des cryotubes congel\u00e9s. Son laboratoire vous enverra un e-mail lorsque la culture aura \u00e9t\u00e9 exp\u00e9di\u00e9e, avec les informations d'exp\u00e9dition et de suivi. Les chercheurs inexp\u00e9riment\u00e9s sont invit\u00e9s \u00e0 suivre des cours sp\u00e9cialis\u00e9s essentiels au travail sur les cellules souches embryonnaires humaines\/iPSC.<\/p>\n

            Le Centre de cellules souches pluripotentes humaines (dirig\u00e9 par le Dr Stanford) de l'Institut de recherche de l'h\u00f4pital d'Ottawa offre une formation individuelle virtuelle sur les bases des techniques de culture des cellules souches pluripotentes humaines (iPSC) sp\u00e9cifiques aux lign\u00e9es cellulaires iPSC de Progeria. Les options et le format de formation sont flexibles en fonction du niveau d'exp\u00e9rience des scientifiques. \u00a0Pour plus d'informations, veuillez envoyer un courriel \u00e0 hpscf@ohri.ca<\/a>.<\/u><\/strong><\/p>\n

            \u00c9tape 4 : L\u2019Universit\u00e9 d\u2019Ottawa vous facturera directement pour chaque ligne iPSC plus les frais de messagerie, le cas \u00e9ch\u00e9ant.<\/strong><\/p>\n

            [\/et_pb_text][et_pb_text admin_label=”mediaprep” module_id=”mediaprep” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

            9. Pr\u00e9paration des milieux de culture cellulaire HGPS et iPS de contr\u00f4le<\/strong><\/h4>\n

            Les cellules iPSC et les cellules souches embryonnaires doivent \u00eatre aliment\u00e9es avec le mTeSR Plus de Stem Cell Technologies (cat# 5825). Veuillez suivre les recommandations du fournisseur pour le stockage.<\/p>\n

            [\/et_pb_text][et_pb_text admin_label=”surfaces” module_id=”surfaces” _builder_version=”4.27.5″ custom_padding=”||0px||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

            10. Pr\u00e9paration des plaques de Matrigel<\/strong><\/h4>\n

            Note<\/strong>:Toutes les \u00e9tapes impliquant le matrigel doivent \u00eatre effectu\u00e9es le plus rapidement possible et rester aussi froides que possible.<\/p>\n

              \n
            1. D\u00e9congeler la bouteille de matrigel \u00e0 4\u00b0<\/strong>C. V\u00e9rifiez le certificat d\u2019analyse de ce lot pour conna\u00eetre sa concentration en prot\u00e9ines.<\/li>\n
            2. Ajoutez suffisamment de milieu DMEM\/F12 froid au matrigel d\u00e9congel\u00e9 pour atteindre une concentration finale de 5 mg\/mL.<\/li>\n
            3. Pr\u00e9parez des aliquotes de 1 ml du matrigel pr\u00e9par\u00e9 \u00e0 l\u2019\u00e9tape 2 dans des tubes Falcon pr\u00e9-refroidis de 15 ml.<\/li>\n
            4. Congeler et conserver tous les aliquots \u00e0 -20\u00b0<\/strong>C.<\/li>\n
            5. Pour fabriquer des plaques de matrigel, pr\u00e9levez une aliquote de matrigel (1 ml) de -20\u00b0<\/strong>C et ajouter 10 ml de DMEM\/F12 froid. Bien m\u00e9langer jusqu'\u00e0 ce que le culot d\u00e9cong\u00e8le (sans cr\u00e9er de bulles et toujours garder la solution froide).<\/li>\n
            6. Transf\u00e9rer dans un tube de 50 ml, puis ajouter 20 ml de DMEM\/F12 froid (1 ml d'aliquote de matrigel est dilu\u00e9 dans 30 ml de DMEM\/F12), bien m\u00e9langer.<\/li>\n
            7. Plaque (1 ml\/puits pour une plaque \u00e0 6 puits, 0,5 ml\/puits pour une plaque \u00e0 12 puits, 0,25 ml\/puits pour une plaque \u00e0 24 puits). Assurez-vous que la solution recouvre toute la surface en secouant doucement la plaque. Ne recongelez pas les restes de matrigel.<\/li>\n
            8. Si vous utilisez la plaque imm\u00e9diatement, laissez-la reposer \u00e0 temp\u00e9rature ambiante pendant 1 heure (ou 30 minutes \u00e0 37\u00b0<\/strong>C), observer le matrigel au microscope. Le matrigel doit \u00eatre bien dispers\u00e9 et non \u00ab grumeleux \u00bb.<\/li>\n
            9. Si vous utilisez les plaques \u00e0 un autre moment, enveloppez le bord de la plaque avec du parafilm et conservez-la \u00e0 4\u00b0<\/strong>C pendant 2 semaines maximum.<\/li>\n<\/ol>\n

              Matrigel \u2013 BD\/Fisher, cat#CB-40230<\/p>\n

              DMEM\/F12 \u2013 Technologies du vivant, cat#11330-057<\/p>\n

              Dr William Stanford-2022<\/p>\n

              [\/et_pb_text][et_pb_text admin_label=”inactivating” module_id=”inactivating” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

              11. <\/strong>D\u00e9cong\u00e9lation des cellules ES ou iPS (par cryo-flacon)<\/strong><\/h4>\n
                \n
              1. Sortez une plaque de matrigel de 4\u00b0C et r\u00e9chauffez-la \u00e0 temp\u00e9rature ambiante pendant une heure, ou pr\u00e9parez une nouvelle plaque de matrigel (voir le protocole de pr\u00e9paration de la plaque de matrigel).<\/li>\n
              2. R\u00e9chauffez 4 ml de mTeSR Plus dans un tube Falcon de 15 ml.<\/li>\n
              3. Retirez les cellules du r\u00e9servoir d'azote liquide et faites-les tourner dans un bain \u00e0 37\u00b0C jusqu'\u00e0 ce qu'il ne reste qu'un petit morceau de glace. Le flacon devrait d\u00e9congeler en 1 \u00e0 2 minutes. Cette \u00e9tape doit \u00eatre effectu\u00e9e rapidement.<\/li>\n
              4. Tube de cellule d'\u00e9thanol et tube de milieu Falcon et placer dans la hotte.<\/li>\n
              5. Utilisez 1 ml pointe de bouche large pour lentement <\/u>ajouter les cellules \u00e0 4 ml de milieu pr\u00e9chauff\u00e9 (\u00e9viter de m\u00e9langer la suspension cellulaire).<\/li>\n
              6. Essorer \u00e0 130 rcf pendant 5 minutes.<\/li>\n
              7. Retirer le surnageant.<\/li>\n
              8. Ajoutez 2 ml de milieu PSC et, avec une pointe \u00e0 large ouverture, briser les mottes doucement<\/u>. Transf\u00e9rer le milieu dans un puits d'une plaque \u00e0 6 puits, ajouter 2 \u00b5l d'inhibiteur ROCK (Y27632, concentration finale de 10 \u00b5M). Placer dans un puits recouvert de matrigel (d'une plaque \u00e0 6 puits).<\/li>\n
              9. Secouez doucement les cellules pour les r\u00e9partir uniform\u00e9ment et placez-les dans un incubateur hypoxique (5%O2<\/sub>, 10% CO2<\/sub>). \u00c9vitez de perturber les plaques pendant 24 heures apr\u00e8s le semis.NOTE: <\/strong>Il est tr\u00e8s important d'\u00e9viter une d\u00e9composition excessive des amas ou un pipetage agressif. Cela peut r\u00e9duire consid\u00e9rablement le taux de survie. Les cellules doivent rester en morceaux de 100 \u00e0 300 cellules au moment de l'ensemencement. Tout en \u00e9tant doux, essayez de travailler rapidement une fois les cellules d\u00e9congel\u00e9es pour minimiser le temps pendant lequel elles sont en contact avec le cryoprotecteur.<\/li>\n
              10. Retirer le milieu apr\u00e8s 24 heures et ajouter 2 ml de milieu PSC (pour une plaque \u00e0 6 puits, 1 ml pour une plaque \u00e0 12 puits et 0,5 ml pour une plaque \u00e0 24 puits). Voir le protocole de r\u00e9colte et d'entretien des hESC\/iPSC.<\/li>\n<\/ol>\n

                M\u00e9dias PSC<\/strong><\/p>\n

                mTeSR Plus de Stem Cell Technologies (cat# 5825). Veuillez suivre les recommandations du fournisseur pour le stockage.<\/p>\n

                Qu'est-ce que l'inhibiteur ROCK Y27632 ?<\/strong><\/p>\n

                L'inhibiteur ROCK Y27632 est un inhibiteur s\u00e9lectif de la kinase p160 ROCK associ\u00e9e \u00e0 Rho. Le traitement avec l'inhibiteur ROCK Y27632 pr\u00e9vient l'apoptose induite par dissociation des cellules souches embryonnaires humaines (hESC) et des cellules souches pluripotentes induites humaines (iPSC), augmentant le taux de survie et maintenant la pluripotence pendant la sous-culture et la d\u00e9cong\u00e9lation des hESC et des hiPSC. Il a \u00e9galement \u00e9t\u00e9 d\u00e9montr\u00e9 que l'inhibiteur ROCK Y27632 am\u00e9liore le taux de survie des cellules souches pendant la cryoconservation. Notez que les aliquotes d'inhibiteur Rock sont sensibles \u00e0 la lumi\u00e8re et aux cycles r\u00e9p\u00e9titifs de cong\u00e9lation-d\u00e9cong\u00e9lation. Assurez-vous d'utiliser ces aliquotes pendant la dur\u00e9e de conservation recommand\u00e9e par le fournisseur.<\/p>\n

                Dr William Stanford-2022<\/p>\n

                [\/et_pb_text][et_pb_text admin_label=”thawing” module_id=”thawing” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

                12. <\/strong>R\u00e9colte et entretien des cellules souches embryonnaires humaines (hESC) et cellules souches embryonnaires in vitro (iPSC)<\/strong><\/h4>\n
                  \n
                1. Le lendemain de la d\u00e9cong\u00e9lation des cellules, examinez-les au microscope pour d\u00e9terminer le taux de survie. REMARQUE : il est normal d'observer un nombre \u00e9lev\u00e9 de cellules non attach\u00e9es. Tant que certaines cellules sont attach\u00e9es, des colonies peuvent en \u00e9merger en 3 \u00e0 7 jours.<\/li>\n
                2. Retirer le milieu des puits et pipeter 2 ml (pour une plaque \u00e0 6 puits, 1 ml pour une plaque \u00e0 12 puits et 0,5 ml pour une plaque \u00e0 24 puits) de milieu PSC frais et chaud par puits. Remettre la plaque dans l'incubateur.<\/li>\n
                3. Les cellules sont nourries jusqu'\u00e0 ce que 60-70% soit confluent (suivre les recommandations du fournisseur).<\/li>\n
                4. \u00c0 partir du deuxi\u00e8me jour, les cellules doivent \u00eatre observ\u00e9es et nettoy\u00e9es de toutes les cellules diff\u00e9renci\u00e9es qui pourraient se d\u00e9velopper.<\/li>\n
                5. Pour nettoyer les cellules, utilisez la hotte de pr\u00e9l\u00e8vement et grattez les cellules diff\u00e9renci\u00e9es avec une pointe de pipette.<\/li>\n
                6. Une fois les cellules nettoy\u00e9es, changez le support comme indiqu\u00e9 dans les \u00e9tapes ci-dessus.<\/li>\n<\/ol>\n

                  (Voir Cong\u00e9lation des hESC\/iPSC ou Transfert des hESC\/iPSC)<\/p>\n

                  NOTE:<\/strong><\/p>\n

                  Il a \u00e9t\u00e9 d\u00e9termin\u00e9 que le milieu PSC \u00e9tait plus efficace dans un environnement hypoxique. Nous avons \u00e9galement observ\u00e9 que la diff\u00e9renciation des cellules \u00e9tait moindre lorsque celles-ci \u00e9taient cultiv\u00e9es dans des incubateurs hypoxiques par rapport \u00e0 des incubateurs normoxiques. Enfin, les cellules ensemenc\u00e9es ont un meilleur taux de survie lorsqu'elles sont cultiv\u00e9es dans un incubateur hypoxique.<\/p>\n

                  Normoxique : 37\u00b0<\/strong>C, 21%O2<\/sub>, 5% CO2<\/sub><\/p>\n

                  Hypoxique : 37\u00b0<\/strong>C, 5%O2<\/sub>, 10% CO2<\/sub><\/p>\n

                  Dr William Stanford-2022<\/p>\n

                  [\/et_pb_text][et_pb_text admin_label=”routine” module_id=”routine” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

                  13. <\/strong>Passage des hESC\/iPSC<\/strong><\/h4>\n
                    \n
                  1. Ajoutez 2 ml de milieu PSC \u00e0 une plaque rev\u00eatue de matrigel \u00e0 6 puits et r\u00e9servez.<\/li>\n
                  2. Prenez la plaque \u00e0 repiquer et retirez le milieu du puits et lavez une fois avec 1 ml de PBS (-\/-).<\/li>\n
                  3. Ajoutez 1 ml de solution EDTA dans le puits et laissez reposer 3 \u00e0 4 minutes \u00e0 temp\u00e9rature ambiante. Ne d\u00e9placez pas la plaque car les cellules peuvent commencer \u00e0 se d\u00e9tacher.<\/li>\n
                  4. Retirez la solution EDTA et ajoutez 1 ml de milieu PSC. Ne laissez pas l'EDTA sur les cellules pendant plus de 4 minutes, car cela provoquerait le d\u00e9collement des cellules.<\/li>\n
                  5. Grattez les cellules \u00e0 l'aide d'un grattoir \u00e0 cellules et r\u00e9partissez-les dans les 6 puits de votre plaque contenant le milieu PSC. \u00c9vitez de briser excessivement les morceaux de la colonie et essayez d'\u00eatre doux avec le grattage. Essayez de conserver les cellules en gros morceaux. Utilisez une pointe de pipette \u00e0 large ouverture pour briser les amas si n\u00e9cessaire. Une fragmentation excessive des cellules peut entra\u00eener la mort cellulaire ou une diff\u00e9renciation spontan\u00e9e excessive apr\u00e8s le passage.<\/li>\n
                  6. Incuber \u00e0 37\u00b0<\/strong>C apr\u00e8s avoir r\u00e9parti uniform\u00e9ment les cellules dans chaque puits (agitation en forme de 8 ou de L). \u00c9viter de perturber la plaque pendant 24 heures apr\u00e8s le passage.<\/li>\n<\/ol>\n

                    NOTE<\/strong>:Une fois les cellules gratt\u00e9es, vous souhaitez les transf\u00e9rer sur la nouvelle plaque d\u00e8s que possible car les cellules se rattacheront rapidement (dans les 5 minutes).<\/p>\n

                    Si l'EDTA reste sur les cellules pendant plus de 4 minutes, les cellules peuvent commencer \u00e0 se d\u00e9tacher. Si cela se produit, r\u00e9cup\u00e9rez simplement les cellules dans un Falcon de 15 ml avec 4 ml de milieu PSC. Faites tourner les cellules \u00e0 130 rcf pendant 5 minutes. Remettez le culot en suspension avec 1 ml de milieu et r\u00e9partissez-le uniform\u00e9ment dans une plaque rev\u00eatue de matrigel \u00e0 6 puits (160 \u00b5l par puits).<\/p>\n

                    Solution EDTA : ajouter 500 \u00b5l d'EDTA 0,5 M (pH 8,0) dans 500 ml de DPBS (-\/-). Ajouter 0,9 g de NaCl. Filtrer la solution pour la st\u00e9riliser et la conserver \u00e0 4 \u00b0C jusqu'\u00e0 6 mois. <\/strong><\/p>\n

                    Extrait du journal :<\/p>\n

                    Passage et expansion de colonies de cellules souches pluripotentes humaines par dissociation sans enzyme dans des conditions de culture chimiquement d\u00e9finies<\/strong><\/p>\n

                    Bi\u00e8res Jeanette<\/a>,1<\/sup> Daniel R. Gulbranson<\/a>,2,3<\/sup> Nicole George<\/a>,4<\/sup>Lauren I. Siniscalchi<\/a>,1<\/sup> Jeffrey Jones<\/a>,4,5<\/sup> James A. Thomson<\/a>,2,3,6<\/sup> et Guokai Chen<\/a>1,2<\/sup><\/p>\n

                    [\/et_pb_text][et_pb_text admin_label=”culturing” module_id=”culturing” _builder_version=”4.27.5″ custom_padding=”||||false|false” z_index_tablet=”500″ global_colors_info=”{}”]<\/p>\n

                    14. <\/strong>Cong\u00e9lation des cellules souches embryonnaires humaines (hESC) et cellules souches embryonnaires in vitro (iPSC) <\/strong><\/h4>\n
                      \n
                    1. Allumez le Bio-Cool (cong\u00e9lateur \u00e0 temp\u00e9rature contr\u00f4l\u00e9e) et r\u00e9glez la temp\u00e9rature \u00e0 -7\u00b0C.<\/li>\n
                    2. Retirez les cellules de l\u2019incubateur et observez la confluence et la morphologie au microscope.<\/li>\n
                    3. Si les puits sont confluents avec 70%, retirez l'ancien milieu et lavez une fois avec du PBS (-\/-), puis ajoutez 1 ml de solution EDTA (voir passage avec la solution EDTA) par puits.<\/li>\n
                    4. Incuber \u00e0 temp\u00e9rature ambiante pendant 3 \u00e0 4 minutes.<\/li>\n
                    5. Aspirez la solution EDTA et ajoutez 1 mL de milieu mFreSR froid (cat#05855, Stem Cell Technologies).<\/li>\n
                    6. Utilisez un grattoir \u00e0 cellules pour soulever d\u00e9licatement les cellules. Conservez les cellules en gros morceaux autant que possible et \u00e9vitez de les pipeter de haut en bas.<\/li>\n
                    7. Transf\u00e9rez les cellules\/mFreSR dans un cryotube \u00e0 l'aide d'un embout \u00e0 large ouverture. Conservez les flacons sur glace jusqu'\u00e0 l'\u00e9tape 8.<\/li>\n
                    8. Placer les tubes dans Bio-Cool et incuber pendant 10 minutes.<\/li>\n
                    9. Obtenez de l'azote liquide.<\/li>\n
                    10. Apr\u00e8s 10 minutes, ensemencez les cellules en trempant une spatule dans de l'azote liquide et en touchant le c\u00f4t\u00e9 du cryo-flacon pendant environ 10 \u00e0 30 secondes ou jusqu'\u00e0 ce que vous voyiez un cristal se former sur le c\u00f4t\u00e9 du cryo-flacon.<\/li>\n
                    11. D\u00e9marrez le programme 1 en appuyant sur le bouton \u00ab PROG \u00bb et parcourez le programme en appuyant \u00e0 nouveau sur le bouton et vous devriez voir le taux de 0,5\u00b0C\/min. , puis appuyez sur \u00ab RUN \u00bb.<\/li>\n
                    12. Une fois la temp\u00e9rature atteinte -65\u00b0C, les cryotubes peuvent \u00eatre transf\u00e9r\u00e9s et stock\u00e9s dans de l'azote liquide.<\/li>\n<\/ol>\n

                      Alternatives<\/p>\n

                      Une autre option consiste \u00e0 placer les cryotubes dans un r\u00e9cipient de cong\u00e9lation (Biocision-CoolCell) et \u00e0 les conserver \u00e0 -80\u00b0C pendant la nuit. Le lendemain, transf\u00e9rez les cryotubes dans l'azote liquide (en phase liquide ou vapeur).<\/p>\n

                      Dr William Stanford-2022<\/p>\n

                      [\/et_pb_text][\/et_pb_column_inner][\/et_pb_row_inner][\/et_pb_column][\/et_pb_section][et_pb_section fb_built=\u201d1\u2033 module_class=\u201dfooter\u201d _builder_version=\u201d4.21.0\u2033 background_color=\u201d#29327a\u201d custom_margin=\u201d-2px|||||\u201d custom_padding=\u201d0|0px|0|0px|false|false\u201d z_index_tablet=\u201d500\u2033 border_width_top=\u201d12px\u201d border_color_top=\u201d#00b2e2\u2033 global_module=\u201d133\u2033 locked=\u201doff\u201d global_colors_info=\u201d{}\u201d][et_pb_row column_structure=\u201d1_4,1_4,1_2\u2033 make_equal=\u201don\u201d module_class=\u201d et_pb_row_fullwidth\u201d _builder_version=\u201d4.16\u2033 width=\u201d89%\u201d width_tablet=\u201d80%\u201d width_phone=\u201d\u201d width_last_edited=\u201don|desktop\u201d max_width=\u201d89%\u201d max_width_tablet=\u201d80%\u201d max_width_phone=\u201d\u201d max_width_last_edited=\u201don|desktop\u201d z_index_tablet=\u201d500\u2033 make_fullwidth=\u201don\u201d width_unit=\u201doff\u201d custom_width_percent=\u201d100%\u201d global_colors_info=\u201d{}\u201d][et_pb_column type=\u201d1_4\u2033 _builder_version=\u201d4.16\u2033 custom_padding=\u201d|||\u201d global_colors_info=\u201d{}\u201d custom_padding__hover=\u201d|||\u201d][et_pb_cta button_url=\u201dhttps:\/\/lp.constantcontactpages.com\/sl\/88gWWwz\u201d button_text=\u201dS'inscrire maintenant\u201d admin_label=\u201dS'inscrire aux mises \u00e0 jour\u201d module_class=\u201dsign-btn\u201d _builder_version=\u201d4.27.4\u2033 header_font_size=\u201d25px\u201d background_color=\u201d#29327a\u201d animation_style=\u201dslide\u201d animation_direction=\u201dleft\u201d animation_intensity_slide=\u201d25%\u201d link_option_url=\u201dhttps:\/\/lp.constantcontactpages.com\/sl\/88gWWwz\u201d header_font_size_tablet=\u201d\u201d header_font_size_phone=\u201d30px\u201d header_font_size_last_edited=\u201don|desktop\u201d z_index_tablet=\u201d500\u2033 border_radii=\u201don|25px|25px|25px|25px\u201d global_colors_info=\u201d{}\u201d button_bg_color__hover_enabled=\u201don\u201d button_bg_color__hover=\u201d#8fd2ed\u201d button_border_color__hover_enabled=\u201don\u201d]<\/p>\n

                      S'inscrire<\/h2>\n

                      pour notre<\/h2>\n

                      Mises \u00e0 jour\u00a0!<\/h2>\n

                      [\/et_pb_cta][\/et_pb_column][et_pb_column type=\u201d1_4\u2033 _builder_version=\u201d4.16\u2033 custom_padding=\u201d|||\u201d global_colors_info=\u201d{}\u201d custom_padding__hover=\u201d|||\u201d][et_pb_cta button_url=\u201dhttps:\/\/progeriaresearch.donorsupport.co\/-\/XZHJVWZR\u201d button_text=\u201dFaites un don maintenant\u201d admin_label=\u201dEnsemble, nous trouverons le rem\u00e8de !\u201d module_class=\u201dsign-btn\u201d _builder_version=\u201d4.16\u2033 header_font_size=\u201d25px\u201d background_color=\u201d#29327a\u201d animation_style=\u201dslide\u201d animation_direction=\u201dleft\u201d animation_intensity_slide=\u201d25%\u201d header_font_size_tablet=\u201d\u201d header_font_size_phone=\u201d30px\u201d header_font_size_last_edited=\u201don|desktop\u201d body_font_size_tablet=\u201d\u201d body_font_size_phone=\u201d\u201d body_font_size_last_edited=\u201don|desktop\u201d z_index_tablet=\u201d500\u2033 border_radii=\u201don|25px|25px|25px|25px\u201d global_colors_info=\u201d{}\u201d button_bg_color__hover_enabled=\u201don\u201d bouton_bg_color__hover=\u201d#8fd2ed\u201d bouton_border_color__hover_enabled=\u201don\u201d]<\/p>\n

                      Ensemble, nous<\/h2>\n

                      VOLONT\u00c9<\/em><\/h2>\n

                      trouve le rem\u00e8de !<\/h2>\n

                      [\/et_pb_cta][\/et_pb_column][et_pb_column type=”1_2″ _builder_version=”4.16″ custom_padding=”|||” global_colors_info=”{}” custom_padding__hover=”|||”][et_pb_image src=”https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2026\/03\/2026-footer-image-copy.png” title_text=”2026 footer image copy” _builder_version=”4.27.5″ _module_preset=”default” custom_margin=”35px||||false|false” global_colors_info=”{}”][\/et_pb_image][\/et_pb_column][\/et_pb_row][\/et_pb_section]<\/p>","protected":false},"excerpt":{"rendered":"

                      [et_pb_section fb_built=\u201d1\u2033 fullwidth=\u201don\u201d disabled_on=\u201doff|off|off\u201d _builder_version=\u201d4.16\u2033 border_width_bottom=\u201d55px\u201d border_color_bottom=\u201d#29327a\u201d locked=\u201doff\u201d global_colors_info=\u201d{}\u201d][et_pb_fullwidth_header _builder_version=\u201d4.16\u2033 title_font=\u201d||||||||\u201d title_font_size=\u201d55\u2033 background_color=\u201d#29327a\u201d background_image=\u201dhttps:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2020\/12\/Jes.jpg\u201d background_position=\u201dcenter_left\u201d custom_padding=\u201d9vw|0px|9vw||true|\u201d custom_padding_tablet=\u201d\u201d custom_padding_phone=\u201d|56px||\u201d custom_padding_last_edited=\u201dsur|bureau\u201d title_font_size_tablet=\u201d45px\u201d title_font_size_phone=\u201d40px\u201d title_font_size_last_edited=\u201dsur|t\u00e9l\u00e9phone\u201d z_index_tablet=\u201d500\u2033 custom_css_main_element=\u201dbackground-position: center 18% !important;\u201d global_colors_info=\u201d{}\u201d] Cellules souches pluripotentes induites [\/et_pb_fullwidth_header][\/et_pb_section][et_pb_section fb_built=\u201d1\u2033 use_custom_gutter=\u201don\u201d gutter_width=\u201d1\u2033 specialty=\u201don\u201d padding_left_1=\u201d35px\u201d padding_left_2=\u201d35px\u201d padding_2_tablet=\u201d|||0px\u201d padding_2_phone=\u201d\u201d padding_2_last_edited=\u201don|desktop\u201d module_class_1=\u201dsidebar-secondary-nav\u201d module_class=\u201dhandprint-bg\u201d _builder_version=\u201d4.16\u2033 background_image=\u201dhttps:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/04\/blue-handprint-only.png\u201d parallax=\u201don\u201d parallax_method=\u201doff\u201d inner_width=\u201d100%\u201d inner_max_width=\u201d100%\u201d custom_padding=\u201d0|0px|54px|0px|false|false\u201d z_index_tablet=\u201d500\u2033 border_width_top=\u201d10px\u201d border_color_top=\u201d#8fd2ed\u201d [\u2026]<\/p>","protected":false},"author":1,"featured_media":0,"parent":0,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"","meta":{"_et_pb_use_builder":"on","_et_pb_old_content":"\t\t\t\t[vc_row][vc_column][vc_custom_heading text=\"Induced Pluripotent Stem Cells\" font_container=\"tag:h1|text_align:center\" use_theme_fonts=\"yes\"][vc_column_text]\r\n\r\n\r\n\r\n
                      \"\"<\/td>\r\n<\/td>\r\n<\/tr>\r\n<\/tbody>\r\n<\/table>\r\nThe Progeria Research Foundation Cell & Tissue Bank\r\n<\/strong>Human Induced Pluripotent Stem Cells (iPSC)\r\n
                        \r\n \t
                      1. Progeria iPSCs <\/strong>Background Information for the Non-scientist<\/strong><\/a>\u00a0<\/strong><\/li>\r\n \t
                      2. Purpose of <\/strong>Induced Pluripotent Stem Cell Generation and Distribution by The Progeria Research Foundation\u00a0<\/strong>\u00a0<\/a><\/li>\r\n \t
                      3. Generation of Hutchinson-Gilford Progeria Syndrome Induced-Pluripotent Stem Cells (iPSCs)<\/strong>\u00a0<\/a><\/li>\r\n \t
                      4. Quality Control: Validation and Characterization<\/strong>\u00a0<\/a><\/li>\r\n \t
                      5. Original Starting Material from which iPSCs Were Derived<\/strong><\/a><\/li>\r\n \t
                      6. Join our Email List for Future iPSC Updates and New Cell Lines<\/strong>\u00a0\u00a0<\/strong><\/a><\/li>\r\n \t
                      7. Questions?\u00a0 Contact us.<\/strong><\/a><\/li>\r\n \t
                      8. Ordering iPSC Lines<\/strong><\/a>\u00a0<\/strong><\/li>\r\n \t
                      9. HGPS and Control iPSC Culture Media Preparation<\/strong>\u00a0<\/strong><\/a><\/li>\r\n \t
                      10. Preparing Madrigal Plates\u00a0<\/strong><\/a><\/li>\r\n \t
                      11. Thawing hESCs and iPSCs<\/strong><\/a><\/li>\r\n \t
                      12. Harvesting and Caring for hESCs and iPSCs<\/strong>\u00a0<\/strong><\/a><\/li>\r\n \t
                      13. Passaging with EDTA solution<\/strong><\/a><\/li>\r\n \t
                      14. Freezing hESCs and iPSCs<\/a><\/strong><\/li>\r\n<\/ol>\r\n\r\n
                        \r\n\r\n                        <\/a>1. iPSC Background information for the non-scientist<\/strong>\r\nStem cells are \u201cimmature\u201d cells that have not yet committed to becoming any one cell type.\u00a0 They are pliable because they have the potential to develop into many different types of mature cells in the body, such as cells that make up the heart or blood vessels, and other tissues and organs.\u00a0 In 2007, researchers discovered a strategy for creating stem cells in the laboratory by reprogramming mature adult cells that we commonly grow for research purposes.1, 2<\/sup><\/strong> . These artificially created stem cells are called Induced Pluripotent Stem Cells (\u201ciPSCs\u201d). For the field of Progeria, this is a huge breakthrough.\u00a0 For the first time, scientists can now make Progeria stem cells and ask questions about how stem cells function and develop in Progeria.\u00a0 Previously there was no source of human Progeria stem cells, and there was therefore a void of information about how Progeria stem cells function compared with stem cells from people without Progeria.\u00a0 In addition, scientists can re-program the Progeria stem cells to create, for the first time, mature Progeria blood vessels, heart cells, and other cell types.\u00a0 Until now, there was no source of human Progeria heart or blood vessel cells.\u00a0 We can now ask key <\/strong>questions about the heart disease that leads to early death in Progeria from heart attacks and strokes. We can compare these discoveries with the heart disease and aging in the general population and discover more about what influences aging in all of us.\u00a0 Already there have been several excellent studies published using Progeria stem cells.3-5<\/sup> \u00a0Our goal at The Progeria Research Foundation is to facilitate many more discoveries using this invaluable tool.\u00a0 For a primer on stem cells, please see this US government website:\u00a0https:\/\/stemcells.nih.gov\/info\/basics.htm<\/a>\r\n
                        \r\n
                          \r\n \t
                        1. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S. Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell. <\/em>2007;131:861-872.<\/li>\r\n \t
                        2. Yu J, Vodyanik MA, Smuga-Otto K, Antosiewicz-Bourget J, Frane JL, Tian S, Nie J, Jonsdottir GA, Ruotti V, Stewart R, Slukvin, II, Thomson JA. Induced pluripotent stem cell lines derived from human somatic cells. Science. <\/em>2007;318:1917-1920.<\/li>\r\n \t
                        3. Liu GH, Barkho BZ, Ruiz S, Diep D, Qu J, Yang SL, Panopoulos AD, Suzuki K, Kurian L, Walsh C, Thompson J, Boue S, Fung HL, Sancho-Martinez I, Zhang K, Yates J, 3rd, Izpisua Belmonte JC. Recapitulation of premature ageing with iPSCs from Hutchinson-Gilford progeria syndrome. Nature. <\/em>2011;472:221-225.<\/li>\r\n \t
                        4. Misteli T. HGPS-derived iPSCs for the ages. Cell Stem Cell. <\/em>2011;8:4-6.<\/li>\r\n<\/ol>\r\n<\/div>\r\n<\/a>2.\u00a0Purpose of induced pluripotent stem cell (iPSC) generation and distribution by The Progeria Research Foundation<\/strong>\r\nThe mission of The Progeria Research Foundation is to discover treatments and the cure for Hutchinson-Gilford Progeria Syndrome and its aging-related disorders. In 2009, PRF entered into a collaboration with an expert team of scientists at the University of Toronto, Canada, under the direction of William Stanford, PhD, to generate high quality Progeria iPSCs. Dr. Stanford is the Canada Research Chair in Integrative Stem Cell Biology. As of 2011, PRF continues to collaborate with Dr. Stanford at the University of Ottawa, Canada where he is Professor of Cellular and Molecular Medicine, Faculty of Medicine, and Senior Scientist at Ottawa Hospital Research Institute's Sprott Centre for Stem Cell Research.<\/a>\r\n\r\nOur goal is to provide this invaluable tool to researchers throughout the world.\u00a0 This new research tool will be used to generate new and innovative research in Progeria, as well as its relationship to heart disease and aging. \u00a0<\/strong>\r\n\r\n3. Generation of Hutchinson-Gilford Progeria Syndrome Induced-Pluripotent Stem Cells (iPSCs)<\/strong><\/a>\r\nInduced-Pluripotent Stem Cells (iPSCs) were derived using VSVG-pseudotyped retroviral transduction of four human factors, Oct4, Sox2, Klf4, and c-Myc into fibroblasts. \u00a0iPSC colonies were derived on mouse-embryonic fibroblasts (MEFs). The procedure used was essentially as previously described but without the use of the EOS reporter (Nature Protocols 4: 1828-1844, 2009). \u00a0<\/strong>\r\n\r\n4. Quality Control: Validation and Characterization<\/strong>\r\nThe lines that are currently available have undergone several validation steps (see downloadable PDFs below):\r\n
                          \r\n
                            \r\n \t
                          1. Mycoplasma Testing for each line: Dr. Stanford\u2019s lab has performed mycoplasmaanalysis by PCR for each cell line.\u00a0 In addition, after expansion and prior to shipping cells, the lines will be retested for mycoplasma.<\/li>\r\n \t
                          2. Immunostaining for pluripotency markers Tra-1-60, Tra-1-81, and SSEA4.<\/li>\r\n \t
                          3. Alkaline Phosphatase Staining as an indicator of pluripotency<\/li>\r\n \t
                          4. Embryoid body formation and subsequent immunostaining for markers of the three germ-layers. Markers tested were \u03b2III-Tubulin (Ectoderm), Smooth-Muscle Actin (Mesoderm), and Gata4 or AFP (Endoderm)<\/li>\r\n \t
                          5. Karyotype analysis.<\/li>\r\n \t
                          6. Re-expression of lamin A in differentiated cells<\/li>\r\n \t
                          7. Teratoma assays<\/li>\r\n<\/ol>\r\n<\/div>\r\n<\/a>\r\nAdditional validation in process:<\/strong>\r\nSome lines have completed teratoma assays as shown in supporting data. For all other lines, teratoma assays are in process and status will be updated as these assays are completed.\r\n\r\n5.\u00a0\u00a0 Original starting material from which these iPS cells were derived<\/strong>\r\niPSCs were derived from PRF Cell & Tissue Bank non-transformed fibroblast cell lines.\r\n\r\nThe transduction method used for all iPS lines was Retrovirus MKOS.\r\n\r\n\r\n\r\n\r\n\r\n\r\n\r\n\r\n\r\n\r\n\r\n\r\n
                            \r\n
                            iPSC Line<\/strong> ID<\/strong>\r\n<\/strong><\/div><\/td>\r\n
                            		\r\n
                            Mutation<\/strong>\r\n<\/strong><\/div><\/td>\r\n
                            		\r\n
                            Gender<\/strong> and Donation<\/strong> Age\r\n<\/strong><\/div><\/td>\r\n
                            		\r\n\r\n
                            Supporting Data\u00a0\"\"<\/strong><\/div><\/td>\r\n<\/tr>\r\n
                            \r\n
                            HGADFN003 iPS 1B<\/div><\/td>\r\n
                            		\r\n
                            \u00a0LMNA<\/em>Exon 11,\r\n1824 C>T<\/div><\/td>\r\n
                            		\r\n
                            Male 2yr 0mo<\/div><\/td>\r\n
                            		\r\n
                            \u00a0<\/strong>Dermal Fibroblasts\r\nHGADFN003<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGADFN003 iPS 1C<\/div><\/td>\r\n
                            		\r\n
                            \u00a0LMNA<\/em> Exon 11,\r\n1824 C>T<\/div><\/td>\r\n
                            		\r\n
                            Male 2yr 0mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts\r\nHGADFN003<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGDFN003\r\niPS 1D<\/div><\/td>\r\n
                            		\r\n
                            \u00a0LMNA <\/em>Exon 11,\r\n1824 C>T<\/div><\/td>\r\n
                            		\r\n
                            Male 2yr 0mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts\r\nHGADFN003<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGADFN167 iPS 1J<\/div><\/td>\r\n
                            		\r\n
                            \u00a0LMNA <\/em>Exon 11, 1824 C>T<\/div><\/td>\r\n
                            		\r\n
                            Male 8yr 5mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts HGADFN167<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGADFN167 iPS 1Q<\/div><\/td>\r\n
                            		\r\n
                            \u00a0LMNA<\/em> Exon 11, 1824 C>T<\/div><\/td>\r\n
                            		\r\n
                            Male 8yr 5mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts HGADFN167<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGMDFN090 iPS 1B<\/div><\/td>\r\n
                            		\r\n
                            \u00a0<\/em>Mother of HGADFN167 (unaffected)<\/div><\/td>\r\n
                            		\r\n
                            Female 37yr 10mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts\r\nHGMDFN090<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGMDFN090 iPS 1C<\/div><\/td>\r\n
                            		\r\n
                            \u00a0<\/em>Mother of HGADFN167 (unaffected)<\/div><\/td>\r\n
                            		\r\n
                            Female 37yr 10mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts\r\nHGMDFN090<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGFDFN168 iPS1 D2<\/div><\/td>\r\n
                            		\r\n
                            \u00a0Father of HGADFN167 (unaffected)<\/div><\/td>\r\n
                            		\r\n
                            Male 40yr\r\n5mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts HGFDFN168<\/div><\/td>\r\n
                            		\r\n
                            \r\n
                            HGFDFN168 iPS1P<\/div><\/td>\r\n
                            		\r\n
                            Father of HGADFN167 (unaffected)<\/div><\/td>\r\n
                            		\r\n
                            Male 40yr<\/div>\r\n
                            5mo<\/div><\/td>\r\n
                            		\r\n
                            Dermal Fibroblasts\r\nHGFDFN168<\/div><\/td>\r\n
                            		<\/a>\r\n
                            168 iPS1P<\/a><\/div><\/td>\r\n<\/tr>\r\n<\/tbody>\r\n<\/table>\r\n
                            <\/div>\r\n\"\"<\/a>\u00a0PRF AVAILABLE CELL LINES<\/a><\/strong>\u00a0<\/strong>\r\n\r\n6.\u00a0Join our email list for future iPSC updates and new cell lines<\/strong>\r\nWe are continuing to generate iPSC lines.\u00a0 If you would like periodic updates on iPSCs held in the PRF Cell & Tissue Bank,\u00a0please join our emailing list by clicking here<\/a><\/strong>\r\n\r\n7.\u00a0Questions?<\/strong><\/a>\r\nPlease contact Leslie Gordon, MD, PhD, Medical Director, with any questions or needs, at lgordon@progeriaresearch.org<\/a> or 978-535-2594\r\n<\/strong>\r\n\r\n8.\u00a0 Ordering iPS cell lines<\/strong>\r\n\r\nIn 2014, PRF instituted a policy of no changes to our MTA. This is the result of 12 years of contractual arrangements with 70 research teams working at institutions in 14 countries. PRF and its counsel have taken into consideration the issues that have arisen in that time period and edited the agreement accordingly, resulting in what we feel are fair and reasonable terms.\r\n\r\nFor U.S. Federal Government Institutions or questions, please contact<\/span>\u00a0<\/span>Wendy Norris at:\u00a0<\/span>wnorris@lifespan.org<\/a><\/span>\u00a0or\u00a0401-274-1122 x 48063<\/a>.\r\n\r\nStep 1: Complete an application and material transfer agreement\r\n<\/strong>\r\n<\/a>\u00a0 Application and Agreement for Non-government Institutions<\/a>\r\n\r\n\u00a0Material Transfer Agreement for Non-government Institutions*<\/a>\r\n\r\nStep 2: Return the completed application and material transfer agreement to PRF at info@progeriaresearch.org<\/a>.\u00a0 Once approved, you will receive an email confirming your order and anticipated shipping date.\u00a0<\/strong><\/a>\r\n\r\nStep 3: <\/strong>Dr. Stanford\u2019s laboratory is currently distributing lines\u00a0in frozen cryovials.\u00a0 His laboratory will email you when the culture has been shipped, with shipping and tracking information. Inexperienced researchers are directed to obtain training at specialized courses essential to human embryonic stem cell\/iPSCs work.<\/a>\r\n\r\nStep 4: The University of Ottawa will charge $84.00 per iPSC line plus courier costs, if any, and will send you a bill directly.<\/strong>\r\n\r\n9.\u00a0 HGPS and Control iPS Cell Culture Media Preparation<\/strong>\r\niPSC and ESCs need to be fed with either TeSR-E8 from Stem Cell\u00a0Technologies (cat# 05990) or Essential 8 medium from ThermoFisher Scientific (cat#A1517001). Please follow the supplier's recommendations for storage.\r\n\r\n10.Preparing Madrigal Plates<\/strong>\r\n\r\nNote<\/strong>: All steps involving matrigel should be done as quickly as possible and stay as cold as possible.\r\n
                              \r\n \t
                            1. Thaw matrigel bottle (10ml) at 4C.<\/li>\r\n \t
                            2. Take matrigel bottle and add 10ml of cold DMEM\/F12 media.<\/li>\r\n \t
                            3. Aliquot 1ml of matrigel in pre-chilled 15ml falcon tubes.<\/li>\r\n \t
                            4. Freeze all aliquots at -20C<\/li>\r\n \t
                            5. To make matrigel plate, remove one aliquot of matrigel (1ml) from -20C and add 18ml of cold DMEM\/F12. Let pellet thaw and once thawed mix well (without creating bubbles) and plate (1ml\/well for 6 well plate, 0.5ml\/well for 12 well plate, 0.25ml\/well for 24 well plate). These plates are labelled as MG1.<\/li>\r\n \t
                            6. If using plate immediately, let plate sit at room temperature for 1 hour, observe matrigel under the microscope. Matrigel should be well dispersed and not \u201cclumpy\u201d.<\/li>\r\n \t
                            7. Once 1 hour is passed transfer matrigel to a second plate (this plate is labelled MG2). If using right away, incubate at room temperature for 1 hour.<\/li>\r\n \t
                            8. If using plates at another time, wrap edge of plate with paraffin and store at 4 degrees.<\/li>\r\n<\/ol>\r\n<\/a>\r\nMatrigel \u2013 BD\/Fisher, cat#CB-40230\r\nDMEM\/F12 \u2013 Life Technologies, cat#11330-057\r\n\r\nDr. William Stanford-2018\r\n\r\n11. Thawing hESCs and iPSCs<\/strong>\r\n\r\nThawing ES or iPS cells (per cryo-vial)<\/u><\/strong>\r\n
                                \r\n \t
                              1. Take matrigel plate out of 4C and warm at room temperature for one hour or make fresh matrigel plate (see preparing matrigel plate protocol<\/strong>).<\/li>\r\n \t
                              2. Warm 4ml of PSC media (pluripotent stem cell media) in a 15ml falcon tube.<\/li>\r\n \t
                              3. Remove cells from freezer and swirl in 37 degree water bath until only a small ice chunk is left. This step must be done quickly.<\/li>\r\n \t
                              4. Ethanol cell tube and falcon media tube and place in the hood.<\/li>\r\n \t
                              5. Use a 1ml wide mouth tip to slowly add cells to 4ml of prewarmed media<\/li>\r\n \t
                              6. Spin at 850rpm for 5min<\/li>\r\n \t
                              7. Remove supernatant<\/li>\r\n \t
                              8. Add 2ml of PSC media and with a\u00a0wide mouth tip break up clumps. Transfer media to one well of a 6 well plate add\u00a02ul of\u00a0ROCK inhibitor (Y27632, final concentration of 10uM)\u200b.<\/li>\r\n \t
                              9. Remove media after 24 hours and add 2ml of PSC media (for a 6 well, 1ml for 12 well and 0.5ml for 24 well plate). See Harvesting and Caring for hESC\/iPSC <\/strong>protocol<\/li>\r\n<\/ol>\r\nPSC media\r\n<\/u><\/strong>TeSR-E8 from Stem Cell Technologies, catalogue # 05990 or Essential 8 Medium from ThermoFisher Scientific, catalogue # A1517001\r\n\r\nWhat is ROCK Inhibitor Y27632?<\/a>\r\n<\/u><\/strong>ROCK Inhibitor Y27632 is a selective inhibitor of the Rhoassociated kinase p160ROCK. Treatment with ROCK Inhibitor Y27632 prevents dissociationinduced apoptosis of human embryonic stem cells (hESC) and human induced pluripotent stem cells (hiPSC), increasing the survival rate and maintaining pluripotency during subcultivation and thawing of hESCs and hiPSCs. ROCK Inhibitor Y27632 also has been shown to enhance the survival rate of stem cells during cryopreservation.\r\n\r\nDr. William Stanford-2018\r\n\r\n12.Harvesting and Caring for hESCs and iPSCs<\/strong>\r\n
                                  \r\n \t
                                1. Remove media from 4C and place in 37C water bath.<\/li>\r\n \t
                                2. The day after cells were thawed take a look at them under the microscope to determine survival rate. (If cells were frozen only with mFreSR the survival rate is appr. 20%, if cells were frozen using the Bio-Cool survival rate is appr. 60%).<\/li>\r\n \t
                                3. Remove media from the wells and pipette 2ml (for a 6 well, 1ml for 12 well and 0.5ml for 24 wells plate) of fresh PSC media per well.<\/li>\r\n \t
                                4. Place plate in 37C incubator (hypoxic condition: 10% CO2 and 5% O2).<\/li>\r\n \t
                                5. Cells are fed every day for the next 3-5 days until 80% confluent.<\/li>\r\n \t
                                6. Cells should be observed and cleaned of any differentiated cells that might be growing.<\/li>\r\n \t
                                7. To clean cells use the picking hood and \u201cscrap off\u201d differentiated cells with a pipette tip.<\/li>\r\n \t
                                8. Once cells are cleaned changed media as done in above steps.<\/li>\r\n<\/ol>\r\n(See Freezing hESC\/iPSC or Passing hESC\/iPSC<\/strong>)\r\n\r\nNOTE:\r\n<\/u><\/strong>PSC media was determine to be more efficient\u00a0 in a hypoxic environment. We have also observed that less differentiation occurs when cells are grown in hypoxic incubators versus normoxic. Also when seeding cells there\u2019s a better survival rate when using a hypoxic incubator.<\/a>\r\n\r\nNormoxic: 5% CO2\r\nHypoxic: 5%O2, 10% CO2\r\n\r\nDr. William Stanford-2018\r\n\r\n13.Passaging with EDTA solution<\/strong>\r\n
                                    \r\n \t
                                  1. Add 2ml of PSC media to a 6 well matrigel coated plate and set aside.<\/li>\r\n \t
                                  2. Take the plate to be passaged and remove the media from the well and wash once with 1ml of PBS(-\/-).<\/li>\r\n \t
                                  3. Add 1ml of\u00a0 the EDTA solution to the well and leave for 3-4min at room temperature. \u00a0Don\u2019t move the plate around as cells can start detaching.<\/li>\r\n \t
                                  4. Once 4 min. is up remove EDTA solution and add 1ml of PSC media. Do not leave EDTA on the cells for more than 4 min as this will cause the cells to lift off.<\/li>\r\n \t
                                  5. Scrape cells (with cell scrapper) and divide cells amongst the 6 wells of your plate containing PSC media (I\u2019ve been taking 160ul into each well). Avoid breaking up the pieces in very small pieces. Try to keep large chunks. Preferably use a wide mouth pipette tip.<\/li>\r\n<\/ol>\r\nNOTE<\/u><\/strong>: Once the cells have been scraped you want to transfer them to the new plate as soon as possible because the cells will reattach quickly (within 5min).\r\n\r\nIf EDTA is left on the cells for more than 4mins. the cells can start to detach. If this happen simply scrap cells and collect in a 15ml falcon with 4ml of PSC media. Spin cells at 850rpm for 5 min. Resuspend the pellet with 1ml of media and divide evenly among a 6 well matrigel coated plate (160ul per well).\r\n\r\nEDTA solution<\/u><\/strong>:\u00a0 Add 500ul of 0.5M EDTA (pH 8.0) into 500ml of DPBS (-\/-). Add 0.9g of NaCl and adjust the osmolarity to 340 mOsm. Filter the solution to sterilize and store it at 4C for up to 6 months. We want the least amount of disturbance for the cells during dissociation therefore the EDTA solution is at the same osmolarity as the PSC media. <\/u><\/strong>\r\n\r\nFrom the paper:<\/a>\r\nPassaging and colony expansion of human pluripotent stem cells by enzyme-free dissociation in chemically defined culture conditions\r\n<\/strong>Jeanette Beers<\/a>,1<\/sup>\u00a0Daniel R. Gulbranson<\/a>,2,3<\/sup>\u00a0Nicole George<\/a>,4<\/sup>Lauren I. Siniscalchi<\/a>,1<\/sup>\u00a0Jeffrey Jones<\/a>,4,5<\/sup>\u00a0James A. Thomson<\/a>,2,3,6<\/sup>\u00a0and\u00a0Guokai Chen<\/a>1,2<\/sup>\r\n\r\n14.Freezing hESCs and iPSCs<\/strong>\r\n
                                      \r\n \t
                                    1. Turn on Bio-Cool (Controlled Rate Freezer) and adjust SP temperature to -7C.<\/li>\r\n \t
                                    2. Remove cells from incubator and observe confluency and morphology under the microscope.\r\nNOTE:<\/strong> Looking for ~80% confluency<\/li>\r\n \t
                                    3. If confluent, remove old media and wash once with PBS(-\/-) then add 1 ml of EDTA solution (see passaging with EDTA solution) per well.<\/li>\r\n \t
                                    4. Incubate at room temperature for 3-4 minutes.<\/li>\r\n \t
                                    5. Aspirate the EDTA solution and add 1 ml of mFreSR media (cat#05855, Stem Cell Technologies)<\/li>\r\n \t
                                    6. Use cell scraper to detach the cells from the well (gently).<\/li>\r\n \t
                                    7. Transfer cells\/mFreSR to a cryotube using a wide mouth tip.<\/li>\r\n \t
                                    8. Place tubes in Bio-Cool and incubate for 10min.<\/li>\r\n \t
                                    9. Get liquid nitrogen.<\/li>\r\n \t
                                    10. After 10min. seed the cells by dipping a spatula into liquid nitrogen and touching the side of the cryo-vial for approximately 10 seconds or until you see a crystal form on the side of the cryo-vial.<\/li>\r\n \t
                                    11. Start program 1 by pushing \u201cPROG\u201d button and going through the program by pushing the button again and you should see the rate of 0.5C\/min. , then press \u201cRUN\u201d.<\/li>\r\n \t
                                    12. Once temperature reaches -65C the cryo-tubes can be transferred and stored in liquid nitrogen.<\/li>\r\n<\/ol>\r\nAlternatives\r\n<\/u><\/strong>1-Another option is to place the cryo-tubes in a freezing container (Biocision-CoolCell) and store at -80C overnight. The next day store cryo-tubes in liquid nitrogen.\r\n\r\nDr. William Stanford-2018[\/vc_column_text][\/vc_column][\/vc_row]\t\t","_et_gb_content_width":"","footnotes":"","_links_to":"","_links_to_target":""},"class_list":["post-1010","page","type-page","status-publish","hentry"],"yoast_head":"\nInduced | The Progeria Research Foundation<\/title>\n<meta name=\"description\" content=\"To learn more about Induced Pluripotent Stem Cells, please contact Leslie Gordon, MD, PhD, Medical Director at lgordon@progeriaresearch.org.\" \/>\n<meta name=\"robots\" content=\"index, follow, max-snippet:-1, max-image-preview:large, max-video-preview:-1\" \/>\n<link rel=\"canonical\" href=\"https:\/\/www.progeriaresearch.org\/fr\/induced-pluripotent-stem-cells\/\" \/>\n<meta property=\"og:locale\" content=\"fr_FR\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"Induced | The Progeria Research Foundation\" \/>\n<meta property=\"og:description\" content=\"To learn more about Induced Pluripotent Stem Cells, please contact Leslie Gordon, MD, PhD, Medical Director at lgordon@progeriaresearch.org.\" \/>\n<meta property=\"og:url\" content=\"https:\/\/www.progeriaresearch.org\/fr\/induced-pluripotent-stem-cells\/\" \/>\n<meta property=\"og:site_name\" content=\"The Progeria Research Foundation\" \/>\n<meta property=\"article:publisher\" content=\"https:\/\/www.facebook.com\/ProgeriaResearch\/\" \/>\n<meta property=\"article:modified_time\" content=\"2026-03-17T14:50:58+00:00\" \/>\n<meta property=\"og:image\" content=\"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg\" \/>\n<meta name=\"twitter:card\" content=\"summary_large_image\" \/>\n<meta name=\"twitter:site\" content=\"@Progeria\" \/>\n<meta name=\"twitter:label1\" content=\"Est. reading time\" \/>\n\t<meta name=\"twitter:data1\" content=\"14 minutes\" \/>\n<script type=\"application\/ld+json\" class=\"yoast-schema-graph\">{\"@context\":\"https:\/\/schema.org\",\"@graph\":[{\"@type\":\"WebPage\",\"@id\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/\",\"url\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/\",\"name\":\"Induced | The Progeria Research Foundation\",\"isPartOf\":{\"@id\":\"https:\/\/www.progeriaresearch.org\/ta\/#website\"},\"primaryImageOfPage\":{\"@id\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#primaryimage\"},\"image\":{\"@id\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#primaryimage\"},\"thumbnailUrl\":\"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg\",\"datePublished\":\"2017-02-27T21:26:38+00:00\",\"dateModified\":\"2026-03-17T14:50:58+00:00\",\"description\":\"To learn more about Induced Pluripotent Stem Cells, please contact Leslie Gordon, MD, PhD, Medical Director at lgordon@progeriaresearch.org.\",\"breadcrumb\":{\"@id\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#breadcrumb\"},\"inLanguage\":\"fr-FR\",\"potentialAction\":[{\"@type\":\"ReadAction\",\"target\":[\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/\"]}]},{\"@type\":\"ImageObject\",\"inLanguage\":\"fr-FR\",\"@id\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#primaryimage\",\"url\":\"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg\",\"contentUrl\":\"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg\"},{\"@type\":\"BreadcrumbList\",\"@id\":\"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#breadcrumb\",\"itemListElement\":[{\"@type\":\"ListItem\",\"position\":1,\"name\":\"Home\",\"item\":\"https:\/\/www.progeriaresearch.org\/\"},{\"@type\":\"ListItem\",\"position\":2,\"name\":\"Induced Pluripotent Stem Cells\"}]},{\"@type\":\"WebSite\",\"@id\":\"https:\/\/www.progeriaresearch.org\/ta\/#website\",\"url\":\"https:\/\/www.progeriaresearch.org\/ta\/\",\"name\":\"The Progeria Research Foundation\",\"description\":\"For the Children \u2665 For the Cure\",\"publisher\":{\"@id\":\"https:\/\/www.progeriaresearch.org\/ta\/#organization\"},\"potentialAction\":[{\"@type\":\"SearchAction\",\"target\":{\"@type\":\"EntryPoint\",\"urlTemplate\":\"https:\/\/www.progeriaresearch.org\/ta\/?s={search_term_string}\"},\"query-input\":{\"@type\":\"PropertyValueSpecification\",\"valueRequired\":true,\"valueName\":\"search_term_string\"}}],\"inLanguage\":\"fr-FR\"},{\"@type\":\"Organization\",\"@id\":\"https:\/\/www.progeriaresearch.org\/ta\/#organization\",\"name\":\"The Progeria Research Foundation\",\"url\":\"https:\/\/www.progeriaresearch.org\/ta\/\",\"logo\":{\"@type\":\"ImageObject\",\"inLanguage\":\"fr-FR\",\"@id\":\"https:\/\/www.progeriaresearch.org\/ta\/#\/schema\/logo\/image\/\",\"url\":\"https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/05\/PRF_Logo_2019_optimized.png\",\"contentUrl\":\"https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/05\/PRF_Logo_2019_optimized.png\",\"width\":300,\"height\":86,\"caption\":\"The Progeria Research Foundation\"},\"image\":{\"@id\":\"https:\/\/www.progeriaresearch.org\/ta\/#\/schema\/logo\/image\/\"},\"sameAs\":[\"https:\/\/www.facebook.com\/ProgeriaResearch\/\",\"https:\/\/x.com\/Progeria\",\"https:\/\/www.instagram.com\/progeriaresearch\/\"]}]}<\/script>\n<!-- \/ Yoast SEO plugin. -->","yoast_head_json":{"title":"Induced | The Progeria Research Foundation","description":"To learn more about Induced Pluripotent Stem Cells, please contact Leslie Gordon, MD, PhD, Medical Director at lgordon@progeriaresearch.org.","robots":{"index":"index","follow":"follow","max-snippet":"max-snippet:-1","max-image-preview":"max-image-preview:large","max-video-preview":"max-video-preview:-1"},"canonical":"https:\/\/www.progeriaresearch.org\/fr\/induced-pluripotent-stem-cells\/","og_locale":"fr_FR","og_type":"article","og_title":"Induced | The Progeria Research Foundation","og_description":"To learn more about Induced Pluripotent Stem Cells, please contact Leslie Gordon, MD, PhD, Medical Director at lgordon@progeriaresearch.org.","og_url":"https:\/\/www.progeriaresearch.org\/fr\/induced-pluripotent-stem-cells\/","og_site_name":"The Progeria Research Foundation","article_publisher":"https:\/\/www.facebook.com\/ProgeriaResearch\/","article_modified_time":"2026-03-17T14:50:58+00:00","og_image":[{"url":"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg","type":"","width":"","height":""}],"twitter_card":"summary_large_image","twitter_site":"@Progeria","twitter_misc":{"Est. reading time":"14 minutes"},"schema":{"@context":"https:\/\/schema.org","@graph":[{"@type":"WebPage","@id":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/","url":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/","name":"Induced | The Progeria Research Foundation","isPartOf":{"@id":"https:\/\/www.progeriaresearch.org\/ta\/#website"},"primaryImageOfPage":{"@id":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#primaryimage"},"image":{"@id":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#primaryimage"},"thumbnailUrl":"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg","datePublished":"2017-02-27T21:26:38+00:00","dateModified":"2026-03-17T14:50:58+00:00","description":"To learn more about Induced Pluripotent Stem Cells, please contact Leslie Gordon, MD, PhD, Medical Director at lgordon@progeriaresearch.org.","breadcrumb":{"@id":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#breadcrumb"},"inLanguage":"fr-FR","potentialAction":[{"@type":"ReadAction","target":["https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/"]}]},{"@type":"ImageObject","inLanguage":"fr-FR","@id":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#primaryimage","url":"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg","contentUrl":"https:\/\/www.progeriaresearch.org\/assets\/images\/medical_images\/ottowa(1).jpg"},{"@type":"BreadcrumbList","@id":"https:\/\/www.progeriaresearch.org\/induced-pluripotent-stem-cells\/#breadcrumb","itemListElement":[{"@type":"ListItem","position":1,"name":"Home","item":"https:\/\/www.progeriaresearch.org\/"},{"@type":"ListItem","position":2,"name":"Induced Pluripotent Stem Cells"}]},{"@type":"WebSite","@id":"https:\/\/www.progeriaresearch.org\/ta\/#website","url":"https:\/\/www.progeriaresearch.org\/ta\/","name":"La Fondation de recherche sur la prog\u00e9ria","description":"Pour les enfants \u2665 Pour la gu\u00e9rison","publisher":{"@id":"https:\/\/www.progeriaresearch.org\/ta\/#organization"},"potentialAction":[{"@type":"SearchAction","target":{"@type":"EntryPoint","urlTemplate":"https:\/\/www.progeriaresearch.org\/ta\/?s={search_term_string}"},"query-input":{"@type":"PropertyValueSpecification","valueRequired":true,"valueName":"search_term_string"}}],"inLanguage":"fr-FR"},{"@type":"Organization","@id":"https:\/\/www.progeriaresearch.org\/ta\/#organization","name":"La Fondation de recherche sur la prog\u00e9ria","url":"https:\/\/www.progeriaresearch.org\/ta\/","logo":{"@type":"ImageObject","inLanguage":"fr-FR","@id":"https:\/\/www.progeriaresearch.org\/ta\/#\/schema\/logo\/image\/","url":"https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/05\/PRF_Logo_2019_optimized.png","contentUrl":"https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/05\/PRF_Logo_2019_optimized.png","width":300,"height":86,"caption":"The Progeria Research Foundation"},"image":{"@id":"https:\/\/www.progeriaresearch.org\/ta\/#\/schema\/logo\/image\/"},"sameAs":["https:\/\/www.facebook.com\/ProgeriaResearch\/","https:\/\/x.com\/Progeria","https:\/\/www.instagram.com\/progeriaresearch\/"]}]}},"_links":{"self":[{"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/pages\/1010","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/comments?post=1010"}],"version-history":[{"count":3,"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/pages\/1010\/revisions"}],"predecessor-version":[{"id":21463,"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/pages\/1010\/revisions\/21463"}],"wp:attachment":[{"href":"https:\/\/www.progeriaresearch.org\/fr\/wp-json\/wp\/v2\/media?parent=1010"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}