{"id":1004,"date":"2017-02-27T21:19:33","date_gmt":"2017-02-27T21:19:33","guid":{"rendered":"https:\/\/www.progeriaresearch.org\/?page_id=1004"},"modified":"2025-02-11T09:34:37","modified_gmt":"2025-02-11T14:34:37","slug":"lymphoblast-cell-culture-protocols","status":"publish","type":"page","link":"https:\/\/www.progeriaresearch.org\/ta\/lymphoblast-cell-culture-protocols\/","title":{"rendered":"\u0bb2\u0bbf\u0bae\u0bcd\u0baa\u0bcb\u0baa\u0bbf\u0bb3\u0bbe\u0bb8\u0bcd\u0b9f\u0bcd \u0b9a\u0bc6\u0bb2\u0bcd \u0b95\u0bb2\u0bbe\u0b9a\u0bcd\u0b9a\u0bbe\u0bb0 \u0ba8\u0bc6\u0bb1\u0bbf\u0bae\u0bc1\u0bb1\u0bc8\u0b95\u0bb3\u0bcd"},"content":{"rendered":"<p>[et_pb_section fb_built=&#8221;1&#8243; fullwidth=&#8221;on&#8221; disabled_on=&#8221;off|off|off&#8221; _builder_version=&#8221;4.16&#8243; border_width_bottom=&#8221;55px&#8221; border_color_bottom=&#8221;#29327a&#8221; locked=&#8221;off&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_fullwidth_header _builder_version=&#8221;4.16&#8243; title_font=&#8221;||||||||&#8221; title_font_size=&#8221;55&#8243; background_color=&#8221;#29327a&#8221; background_image=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2020\/12\/Jes.jpg&#8221; background_position=&#8221;center_left&#8221; custom_padding=&#8221;9vw||9vw||true&#8221; custom_padding_tablet=&#8221;&#8221; custom_padding_phone=&#8221;|56px||&#8221; custom_padding_last_edited=&#8221;on|desktop&#8221; title_font_size_tablet=&#8221;45px&#8221; title_font_size_phone=&#8221;40px&#8221; title_font_size_last_edited=&#8221;on|phone&#8221; z_index_tablet=&#8221;500&#8243; custom_css_main_element=&#8221;background-position: center 18% !important;&#8221; global_colors_info=&#8221;{}&#8221;]<\/p>\n<h1 class=\"vc_custom_heading\" data-fontsize=\"34\" data-lineheight=\"48\" style=\"text-align: right;\">Lymphoblast Cell<\/h1>\n<h1 class=\"vc_custom_heading\" data-fontsize=\"34\" data-lineheight=\"48\" style=\"text-align: right;\">Culture Protocols<\/h1>\n<p>&nbsp;<\/p>\n<p>[\/et_pb_fullwidth_header][\/et_pb_section][et_pb_section fb_built=&#8221;1&#8243; use_custom_gutter=&#8221;on&#8221; gutter_width=&#8221;1&#8243; specialty=&#8221;on&#8221; padding_left_1=&#8221;35px&#8221; padding_left_2=&#8221;35px&#8221; padding_2_tablet=&#8221;|||0px&#8221; padding_2_phone=&#8221;&#8221; padding_2_last_edited=&#8221;on|desktop&#8221; module_class_1=&#8221;sidebar-secondary-nav&#8221; module_class=&#8221;handprint-bg&#8221; _builder_version=&#8221;4.16&#8243; background_image=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/04\/blue-handprint-only.png&#8221; parallax=&#8221;on&#8221; parallax_method=&#8221;off&#8221; inner_width=&#8221;100%&#8221; inner_max_width=&#8221;100%&#8221; custom_padding=&#8221;0|0px|54px|0px|false|false&#8221; z_index_tablet=&#8221;500&#8243; border_width_top=&#8221;10px&#8221; border_color_top=&#8221;#8fd2ed&#8221; use_custom_width=&#8221;on&#8221; width_unit=&#8221;off&#8221; custom_width_percent=&#8221;100%&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column type=&#8221;1_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_sidebar area=&#8221;et_pb_widget_area_14&#8243; disabled_on=&#8221;on|on|off&#8221; module_class=&#8221;subpage-sidebars&#8221; _builder_version=&#8221;4.16&#8243; animation_style=&#8221;fade&#8221; z_index_tablet=&#8221;500&#8243; border_width_right=&#8221;5px&#8221; locked=&#8221;off&#8221; global_colors_info=&#8221;{}&#8221;]<br \/>\n[\/et_pb_sidebar][\/et_pb_column][et_pb_column type=&#8221;3_4&#8243; specialty_columns=&#8221;3&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_row_inner _builder_version=&#8221;4.16&#8243; background_color=&#8221;#00b2e2&#8243; custom_margin=&#8221;41px||&#8221; custom_margin_tablet=&#8221;0px||&#8221; custom_margin_phone=&#8221;&#8221; custom_margin_last_edited=&#8221;on|phone&#8221; custom_padding=&#8221;39.4375px|20px|0|20px|false|true&#8221; animation_style=&#8221;slide&#8221; animation_direction=&#8221;top&#8221; animation_intensity_slide=&#8221;25%&#8221; box_shadow_style=&#8221;preset1&#8243; box_shadow_blur=&#8221;38px&#8221; box_shadow_spread=&#8221;-12px&#8221; locked=&#8221;off&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column_inner saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_text admin_label=&#8221;The Progeria Research Foundation\u2019s work&#8230;&#8221; _builder_version=&#8221;4.16&#8243; header_2_line_height=&#8221;1.2em&#8221; background_layout=&#8221;dark&#8221; custom_margin=&#8221;||35px&#8221; custom_padding=&#8221;|50px||&#8221; custom_padding_tablet=&#8221;|50px||50px||true&#8221; custom_padding_phone=&#8221;&#8221; custom_padding_last_edited=&#8221;on|desktop&#8221; header_font_size_tablet=&#8221;&#8221; header_font_size_phone=&#8221;&#8221; header_font_size_last_edited=&#8221;on|desktop&#8221; header_2_font_size_tablet=&#8221;25px&#8221; header_2_font_size_phone=&#8221;&#8221; header_2_font_size_last_edited=&#8221;on|tablet&#8221; header_4_font_size_tablet=&#8221;&#8221; header_4_font_size_phone=&#8221;19px&#8221; header_4_font_size_last_edited=&#8221;on|phone&#8221; header_4_line_height_tablet=&#8221;&#8221; header_4_line_height_phone=&#8221;1.2em&#8221; header_4_line_height_last_edited=&#8221;on|phone&#8221; z_index_tablet=&#8221;500&#8243; global_colors_info=&#8221;{}&#8221;]<\/p>\n<h2 style=\"text-align: center;\"><strong>Protocol for Culturing Transformed Lymphocytes<\/strong><\/h2>\n<p>[\/et_pb_text][\/et_pb_column_inner][\/et_pb_row_inner][et_pb_row_inner custom_padding_last_edited=&#8221;on|phone&#8221; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|35px|0|0px|false|false&#8221; custom_padding_tablet=&#8221;|35px||35px||true&#8221; custom_padding_phone=&#8221;&#8221; animation_direction=&#8221;top&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column_inner saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_text _builder_version=&#8221;4.27.4&#8243; background_size=&#8221;initial&#8221; background_position=&#8221;top_left&#8221; background_repeat=&#8221;repeat&#8221; custom_padding=&#8221;||0px||false|false&#8221; hover_enabled=&#8221;0&#8243; global_colors_info=&#8221;{}&#8221; sticky_enabled=&#8221;0&#8243;]<\/p>\n<h4 style=\"text-align: center;\"><strong>Protocol for Culturing Lymphoblasts<\/strong><\/h4>\n<h4><strong>Growth Medium<\/strong><\/h4>\n<p>RPMI 1640 &#8211; ThermoFisher # 11875<br \/>15% Fetal Bovine Serum (FBS) \u2013 ThermoFisher #10437-028<br \/>1% (1X) Penicillin-Streptomycin (optional) &#8211; ThermoFisher #15140-122<\/p>\n<h4><strong><\/strong><\/h4>\n<h4><strong>General Notes<br \/><\/strong><\/h4>\n<ul>\n<li>Growth medium should always be equilibrated in the incubator at 37\u00b0C, 5% CO<sub>2<\/sub> for 30 minutes before use<\/li>\n<li>Always use flasks with vented caps<\/li>\n<li>Always incubate the flask in an upright position<\/li>\n<li>No more than 20 ml should be used in a T25 flask<\/li>\n<\/ul>\n<h4><strong><\/strong><\/h4>\n<h4><strong>Thawing Frozen Cells<br \/><\/strong><\/h4>\n<p>Frozen cells will be received in 1 ml aliquots on dry ice.\u00a0 Keep cells frozen until thawed for cell culture or place in liquid nitrogen storage if not starting cultures right away.\u00a0 However, it is recommended that if you are going to store the cells for later use, grow up fresh stocks and freeze multiple ampoules.\u00a0 The cells are cryopreserved in 45% RPMI-1640, 50% FBS, and 5% DMSO (tissue culture grade).<\/p>\n<ol>\n<li style=\"list-style-type: none;\">\n<ol>\n<li>Place 10 ml of RPMI-1640, 15% fetal bovine serum (FBS), \u00b1 antibiotics in a T-25 flask with a vented cap.<\/li>\n<li>Allow the media to equilibrate in a 37\u00b0C humidified incubator with 5% CO<sub>2<\/sub> in air for 30 minutes.\u00a0 Flasks should be incubated in an upright position.<\/li>\n<li>Thaw each ampoule or vial of frozen cells in a 37\u00b0C water bath or a beaker of lukewarm water, one at a time.<\/li>\n<li>Quickly clean the outside of the ampoule or vial with 70% ethanol.\u00a0 If the cells are in a glass ampoule, carefully break the ampoule being sure to protect fingers from broken glass.<\/li>\n<li>Remove the 1 ml of frozen cells with a sterile pipet and place in a T-25 flask with 10 ml of medium.\u00a0 Clearly label flask to identify each cell line.<\/li>\n<li>Place the flask in a 37\u00b0C humidified incubator with 5% CO<sub>2<\/sub> in the air.\u00a0 Flasks should be incubated in an upright position.<\/li>\n<li>After 24 hours, remove 5 ml of medium from the top without disturbing lymphocytes settled on the bottom of the flask and replace with 5 ml of pre-warmed medium.<\/li>\n<li>Proceed with the lymphoblast cell culture protocol.<\/li>\n<\/ol>\n<\/li>\n<\/ol>\n<h4><strong><\/strong><\/h4>\n<h4><strong>Lymphoblast Culture<\/strong><\/h4>\n<ol>\n<li style=\"list-style-type: none;\">\n<ol>\n<li>Three to four days after thawing, count cells. Cells are non-adherent and grow in clumps.\u00a0 Break up clumps by gentle pipetting.<\/li>\n<li>Count cells.<\/li>\n<li>Ideally, cell concentration should not exceed 2 x 10<sup>6<\/sup> cells\/ml.<\/li>\n<li>Depending on cell concentration refeed, split or freeze cells.<\/li>\n<li>Refeed by adding 5-6 ml of equilibrated growth medium.<\/li>\n<li>Seed new cultures at no less than 2 x 10<sup>5<\/sup> cells\/ml.<\/li>\n<li>Cells should be frozen at approximately 5 x 10<sup>6<\/sup> cells\/ml. Follow cell freezing protocol.<\/li>\n<\/ol>\n<\/li>\n<\/ol>\n<h4><strong><\/strong><\/h4>\n<h4><strong>Freezing Lymphoblasts<\/strong><\/h4>\n<ol>\n<li style=\"list-style-type: none;\">\n<ol>\n<li>Cells should be frozen at approximately 5 x 10<sup>6<\/sup> cells\/ml.<\/li>\n<li>Transfer appropriate volume of cells to 15 or 50 ml conical tube.<\/li>\n<li>Centrifuge at 4\u00b0C at 100 x g for 10 minutes.<\/li>\n<li>Remove medium and resuspend in appropriate volume of cold freezing medium.<\/li>\n<li>Transfer 1 ml cells each into cryovial.<\/li>\n<li>Place cryovials in freezing chamber and place chamber in -80\u00b0C freezer overnight.<\/li>\n<li>Transfer cryovials to liquid nitrogen the next day.<\/li>\n<\/ol>\n<\/li>\n<\/ol>\n<p>[\/et_pb_text][\/et_pb_column_inner][\/et_pb_row_inner][et_pb_row_inner _builder_version=&#8221;4.27.4&#8243; _module_preset=&#8221;default&#8221; hover_enabled=&#8221;0&#8243; global_colors_info=&#8221;{}&#8221; sticky_enabled=&#8221;0&#8243; custom_padding=&#8221;24px||24px|||&#8221;][et_pb_column_inner saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.27.4&#8243; _module_preset=&#8221;default&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_button button_url=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2025\/02\/Lymphoblast-protocol-1-22-25.pdf&#8221; button_text=&#8221;Protocol for Culturing Lymphoblasts&#8221; admin_label=&#8221;Protocol for Culturing Lymphoblasts&#8221; _builder_version=&#8221;4.27.4&#8243; _module_preset=&#8221;default&#8221; button_text_color=&#8221;#FFFFFF&#8221; button_bg_color=&#8221;#00B2E2&#8243; button_border_color=&#8221;#00B2E2&#8243; global_colors_info=&#8221;{}&#8221;][\/et_pb_button][\/et_pb_column_inner][\/et_pb_row_inner][et_pb_row_inner custom_padding_last_edited=&#8221;on|phone&#8221; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;||0px|||&#8221; custom_padding_tablet=&#8221;|35px||35px||true&#8221; custom_padding_phone=&#8221;&#8221; border_width_top=&#8221;10px&#8221; border_color_top=&#8221;#8fd2ed&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column_inner saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_text _builder_version=&#8221;4.16&#8243; global_colors_info=&#8221;{}&#8221;]<\/p>\n<h4><strong>For Further Information Please Contact:<\/strong><\/h4>\n<p>[\/et_pb_text][\/et_pb_column_inner][\/et_pb_row_inner][et_pb_row_inner column_structure=&#8221;1_2,1_2&#8243; custom_padding_last_edited=&#8221;on|phone&#8221; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|35px|0|0px|false|false&#8221; custom_padding_tablet=&#8221;|35px||35px||true&#8221; custom_padding_phone=&#8221;&#8221; animation_direction=&#8221;top&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column_inner type=&#8221;1_2&#8243; saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|20px||&#8221; custom_padding_tablet=&#8221;||20px&#8221; custom_padding_phone=&#8221;&#8221; custom_padding_last_edited=&#8221;on|desktop&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_text _builder_version=&#8221;4.19.5&#8243; global_colors_info=&#8221;{}&#8221;]<\/p>\n<h6>Leslie B. Gordon, MD, PhD<\/h6>\n<p>Professor of Pediatrics Research\u00a0Warren Alpert Medical School of Brown University and Department of Pediatrics, Hasbro Children&#8217;s Hospital, Providence, RI\u00a0Department of Anesthesia, Children&#8217;s Hospital Boston and Harvard Medical School, Boston, MA\u00a0Medical Director, The Progeria Research Foundation<\/p>\n<p>Phone: 978-535-2594<br \/>Fax: 508-543-0377<br \/><span> <\/span><a href=\"mailto:lgordon@progeriaresearch.org\" title=\"mailto:lgordon@progeriaresearch.org\">lgordon@progeriaresearch.org<\/a><\/p>\n<p>[\/et_pb_text][\/et_pb_column_inner][et_pb_column_inner type=&#8221;1_2&#8243; saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||20px&#8221; custom_padding_tablet=&#8221;20px|||0px&#8221; custom_padding_phone=&#8221;&#8221; custom_padding_last_edited=&#8221;on|phone&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_text _builder_version=&#8221;4.27.2&#8243; custom_padding=&#8221;||0px|||&#8221; custom_padding_last_edited=&#8221;off|desktop&#8221; global_colors_info=&#8221;{}&#8221;]<\/p>\n<h6><span>Wendy Norris<\/span><\/h6>\n<div><span>PRF Cell and Tissue Bank<br \/>Phone: 401-274-1122 x 48063<br \/><a href=\"mailto:wnorris@brownhealth.org\" title=\"mailto:wnorris@brownhealth.org\" data-outlook-id=\"6ae643af-3829-46f7-8b6a-2ef767034802\">wnorris@brownhealth.org<\/a><br \/><\/span><\/div>\n<p>[\/et_pb_text][\/et_pb_column_inner][\/et_pb_row_inner][\/et_pb_column][\/et_pb_section][et_pb_section fb_built=&#8221;1&#8243; module_class=&#8221;footer&#8221; _builder_version=&#8221;4.21.0&#8243; background_color=&#8221;#29327a&#8221; custom_margin=&#8221;-2px|||||&#8221; custom_padding=&#8221;0|0px|0|0px|false|false&#8221; z_index_tablet=&#8221;500&#8243; border_width_top=&#8221;12px&#8221; border_color_top=&#8221;#00b2e2&#8243; global_module=&#8221;133&#8243; locked=&#8221;off&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_row_inner column_structure=&#8221;1_3,1_3,1_3&#8243; disabled_on=&#8221;on|on|on&#8221; admin_label=&#8221;Row&#8221; _builder_version=&#8221;4.21.0&#8243; _module_preset=&#8221;default&#8221; min_height=&#8221;277px&#8221; custom_margin=&#8221;|20px|30px||false|false&#8221; custom_padding=&#8221;0px|30px|0px||false|false&#8221; disabled=&#8221;on&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column_inner type=&#8221;1_3&#8243; saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.21.0&#8243; _module_preset=&#8221;default&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_image src=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2023\/08\/Startbanner-1.png&#8221; title_text=&#8221;Startbanner&#8221; url=&#8221;https:\/\/www.gravoc.com\/&#8221; align=&#8221;right&#8221; _builder_version=&#8221;4.21.0&#8243; _module_preset=&#8221;default&#8221; custom_margin=&#8221;|-50px||20px|false|false&#8221; global_colors_info=&#8221;{}&#8221;][\/et_pb_image][\/et_pb_column_inner][et_pb_column_inner type=&#8221;1_3&#8243; saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.21.0&#8243; _module_preset=&#8221;default&#8221; global_colors_info=&#8221;{}&#8221;][\/et_pb_column_inner][et_pb_column_inner type=&#8221;1_3&#8243; saved_specialty_column_type=&#8221;3_4&#8243; _builder_version=&#8221;4.21.0&#8243; _module_preset=&#8221;default&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_image src=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2023\/07\/RFRFinish.png&#8221; title_text=&#8221;RFRFinish&#8221; _builder_version=&#8221;4.21.0&#8243; _module_preset=&#8221;default&#8221; custom_margin=&#8221;|20px||-50px|false|false&#8221; global_colors_info=&#8221;{}&#8221;][\/et_pb_image][\/et_pb_column_inner][\/et_pb_row_inner][et_pb_row column_structure=&#8221;1_4,1_4,1_2&#8243; make_equal=&#8221;on&#8221; module_class=&#8221; et_pb_row_fullwidth&#8221; _builder_version=&#8221;4.16&#8243; width=&#8221;89%&#8221; width_tablet=&#8221;80%&#8221; width_phone=&#8221;&#8221; width_last_edited=&#8221;on|desktop&#8221; max_width=&#8221;89%&#8221; max_width_tablet=&#8221;80%&#8221; max_width_phone=&#8221;&#8221; max_width_last_edited=&#8221;on|desktop&#8221; z_index_tablet=&#8221;500&#8243; make_fullwidth=&#8221;on&#8221; width_unit=&#8221;off&#8221; custom_width_percent=&#8221;100%&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_column type=&#8221;1_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_cta button_url=&#8221;https:\/\/lp.constantcontactpages.com\/sl\/88gWWwz&#8221; button_text=&#8221;Sign Up Now&#8221; admin_label=&#8221;Sign Up for Updates&#8221; module_class=&#8221;sign-btn&#8221; _builder_version=&#8221;4.27.4&#8243; header_font_size=&#8221;25px&#8221; background_color=&#8221;#29327a&#8221; animation_style=&#8221;slide&#8221; animation_direction=&#8221;left&#8221; animation_intensity_slide=&#8221;25%&#8221; link_option_url=&#8221;https:\/\/lp.constantcontactpages.com\/sl\/88gWWwz&#8221; header_font_size_tablet=&#8221;&#8221; header_font_size_phone=&#8221;30px&#8221; header_font_size_last_edited=&#8221;on|desktop&#8221; z_index_tablet=&#8221;500&#8243; border_radii=&#8221;on|25px|25px|25px|25px&#8221; global_colors_info=&#8221;{}&#8221; button_bg_color__hover_enabled=&#8221;on&#8221; button_bg_color__hover=&#8221;#8fd2ed&#8221; button_border_color__hover_enabled=&#8221;on&#8221;]<\/p>\n<h2>Sign Up<\/h2>\n<h2>for Our<\/h2>\n<h2>Updates!<\/h2>\n<p>[\/et_pb_cta][\/et_pb_column][et_pb_column type=&#8221;1_4&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_cta button_url=&#8221;https:\/\/progeriaresearch.donorsupport.co\/-\/XZHJVWZR&#8221; button_text=&#8221;Donate Now&#8221; admin_label=&#8221;Together, we will find the cure!&#8221; module_class=&#8221;sign-btn&#8221; _builder_version=&#8221;4.16&#8243; header_font_size=&#8221;25px&#8221; background_color=&#8221;#29327a&#8221; animation_style=&#8221;slide&#8221; animation_direction=&#8221;left&#8221; animation_intensity_slide=&#8221;25%&#8221; header_font_size_tablet=&#8221;&#8221; header_font_size_phone=&#8221;30px&#8221; header_font_size_last_edited=&#8221;on|desktop&#8221; body_font_size_tablet=&#8221;&#8221; body_font_size_phone=&#8221;&#8221; body_font_size_last_edited=&#8221;on|desktop&#8221; z_index_tablet=&#8221;500&#8243; border_radii=&#8221;on|25px|25px|25px|25px&#8221; global_colors_info=&#8221;{}&#8221; button_bg_color__hover_enabled=&#8221;on&#8221; button_bg_color__hover=&#8221;#8fd2ed&#8221; button_border_color__hover_enabled=&#8221;on&#8221;]<\/p>\n<h2>Together, we<\/h2>\n<h2><em>WILL<\/em><\/h2>\n<h2>find the cure!<\/h2>\n<p>[\/et_pb_cta][\/et_pb_column][et_pb_column type=&#8221;1_2&#8243; _builder_version=&#8221;4.16&#8243; custom_padding=&#8221;|||&#8221; global_colors_info=&#8221;{}&#8221; custom_padding__hover=&#8221;|||&#8221;][et_pb_image src=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2024\/08\/2024-strip-footer-strip-copy.png&#8221; title_text=&#8221;2024 strip footer strip copy&#8221; _builder_version=&#8221;4.27.0&#8243; _module_preset=&#8221;default&#8221; custom_margin=&#8221;35px||||false|false&#8221; global_colors_info=&#8221;{}&#8221;][\/et_pb_image][\/et_pb_column][\/et_pb_row][\/et_pb_section]<\/p>\n","protected":false},"excerpt":{"rendered":"<p>[et_pb_section fb_built=&#8221;1&#8243; fullwidth=&#8221;on&#8221; disabled_on=&#8221;off|off|off&#8221; _builder_version=&#8221;4.16&#8243; border_width_bottom=&#8221;55px&#8221; border_color_bottom=&#8221;#29327a&#8221; locked=&#8221;off&#8221; global_colors_info=&#8221;{}&#8221;][et_pb_fullwidth_header _builder_version=&#8221;4.16&#8243; title_font=&#8221;||||||||&#8221; title_font_size=&#8221;55&#8243; background_color=&#8221;#29327a&#8221; background_image=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2020\/12\/Jes.jpg&#8221; background_position=&#8221;center_left&#8221; custom_padding=&#8221;9vw||9vw||true&#8221; custom_padding_tablet=&#8221;&#8221; custom_padding_phone=&#8221;|56px||&#8221; custom_padding_last_edited=&#8221;on|desktop&#8221; title_font_size_tablet=&#8221;45px&#8221; title_font_size_phone=&#8221;40px&#8221; title_font_size_last_edited=&#8221;on|phone&#8221; z_index_tablet=&#8221;500&#8243; custom_css_main_element=&#8221;background-position: center 18% !important;&#8221; global_colors_info=&#8221;{}&#8221;] Lymphoblast Cell Culture Protocols &nbsp; [\/et_pb_fullwidth_header][\/et_pb_section][et_pb_section fb_built=&#8221;1&#8243; use_custom_gutter=&#8221;on&#8221; gutter_width=&#8221;1&#8243; specialty=&#8221;on&#8221; padding_left_1=&#8221;35px&#8221; padding_left_2=&#8221;35px&#8221; padding_2_tablet=&#8221;|||0px&#8221; padding_2_phone=&#8221;&#8221; padding_2_last_edited=&#8221;on|desktop&#8221; module_class_1=&#8221;sidebar-secondary-nav&#8221; module_class=&#8221;handprint-bg&#8221; _builder_version=&#8221;4.16&#8243; background_image=&#8221;https:\/\/www.progeriaresearch.org\/wp-content\/uploads\/2019\/04\/blue-handprint-only.png&#8221; parallax=&#8221;on&#8221; parallax_method=&#8221;off&#8221; inner_width=&#8221;100%&#8221; inner_max_width=&#8221;100%&#8221; custom_padding=&#8221;0|0px|54px|0px|false|false&#8221; z_index_tablet=&#8221;500&#8243; border_width_top=&#8221;10px&#8221; border_color_top=&#8221;#8fd2ed&#8221; [&hellip;]<\/p>\n","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=\"Lymphoblast Cell Culture Protocols\" font_container=\"tag:h1|text_align:center\" use_theme_fonts=\"yes\"][vc_column_text]<strong>Protocol for Culturing Transformed Lymphocytes<\/strong>\r\n\r\nFrom Shipment of Frozen Cells\r\n\r\nFrozen cells will be received in 1 ml aliquots on dry ice. Keep cells frozen until thawed for cell culture or place in liquid nitrogen storage if not starting cultures right away. However, it is recommended that if you are going to store the cells for later use, grow up fresh stocks and freeze multiple ampoules. The cells are cryopreserved in 45% RPMI-1640, 50% fetal calf serum, and 5% DMSO (tissue culture grade).\r\n<ol>\r\n \t<li>Place 10 ml of RPMI-1640, 15% fetal calf serum (FCS), \u00b1antibiotics in a T-25 flask.<\/li>\r\n \t<li>Allow the media to equilibrate in a 37\u00b0C humidified incubator with 5% CO 2in air for 30 minutes.<\/li>\r\n \t<li>Thaw each ampoule of frozen cells in a 37\u00b0C water bath or a beaker of lukewarm water, one at a time.<\/li>\r\n \t<li>Quickly clean the outside of the ampoule off with a sterile isopropanol prep pad, then wrap the prep pad around the ampoule to protect fingers and crack the seal on the ampoule inside a biological safety hood.<\/li>\r\n \t<li>Remove the 1 ml of frozen cells with a sterile glass Pasteur pipet and place in a T-25 flask with 10 ml of medium. Clearly label flask to identify each cell line.<\/li>\r\n \t<li>Place the cells in a 37\u00b0C humidified incubator with 5% CO 2in the air.<\/li>\r\n \t<li>After 24 hours remove 5 ml of medium from the top without disturbing lymphocytes settled on the bottom of the flask and replace with 5 ml of pre-warmed medium.<\/li>\r\n \t<li>Feed the cells with 5 - 6 ml of fresh medium every 3 - 4 days and split into new cultures as required.<\/li>\r\n<\/ol>\r\n<strong>From Shipment of Live Cultures<\/strong>\r\n<ol>\r\n \t<li>Live lymphocyte cultures will be received in T-25 flasks filled to the top with RPMI-1640, 15% FCS, 1% Penicillin-Streptomycin (Gibco). The caps will be tight and sealed with parafilm.<\/li>\r\n \t<li>Remove parafilm.<\/li>\r\n \t<li>Place cultures in a 37\u00b0C humidified incubator with 5% CO2 in air and allow cells to settle out (20 - 30 minutes).<\/li>\r\n \t<li>Using a sterile pipet remove all but 10 - 15 ml of medium and discard.<\/li>\r\n \t<li>Place caps back on flasks loosened slightly to allow gas exchange (Check to make sure cap threads do not have media on them. If so you may want to transfer to a new flask.) Place the cultures back in the incubator with 5% CO 2in air. They should recover in a day or two.<\/li>\r\n \t<li>Feed the cells with fresh medium every 3 - 4 days by resuspending the cell aggregates and removing 50 -60% of the volume and replacing with fresh media. Cells removed from the flask can be used to initiate new cultures as needed.<\/li>\r\n<\/ol>\r\n<strong>For Further Information Please Contact:<\/strong>\r\n\r\nLeslie B. Gordon, MD, PhD\r\nProfessor of Pediatrics Research\u00a0Warren Alpert Medical School of Brown University and Department of Pediatrics, Hasbro Children's Hospital, Providence, RI\u00a0Department of Anesthesia, Children's Hospital Boston and Harvard Medical School, Boston, MA\u00a0Medical Director, The Progeria Research Foundation\r\n\r\nPhone: 978-535-2594\r\nFax: 508-543-0377\r\n<a href=\"mailto:Leslie_Gordon@brown.edu\">Leslie_Gordon@brown.edu<\/a>\r\n<div><span style=\"font-family: Georgia, Palatino;\">Wendy Norris<\/span><\/div>\r\n<div><span style=\"font-family: Georgia, Palatino;\">PRF Cell and Tissue Bank\r\nPhone: 401-274-1122 x 48063\r\n<a href=\"mailto:wnorris@lifespan.org\" target=\"_blank\" rel=\"noopener noreferrer\">wnorris@lifespan.org<\/a><\/span><\/div>\r\n[\/vc_column_text][\/vc_column][\/vc_row]\t\t","_et_gb_content_width":"","footnotes":"","_links_to":"","_links_to_target":""},"class_list":["post-1004","page","type-page","status-publish","hentry"],"yoast_head":"<!-- This site is optimized with the Yoast SEO plugin v27.8 - https:\/\/yoast.com\/product\/yoast-seo-wordpress\/ -->\n<title>Lymphoblast | The Progeria Research Foundation<\/title>\n<meta name=\"description\" content=\"Lymphoblast Cell Culture Protocols are listed within our website, regarding from shipment of frozen cells and live cultures.\" \/>\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\/ta\/lymphoblast-cell-culture-protocols\/\" \/>\n<meta property=\"og:locale\" content=\"ta_IN\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"Lymphoblast | The Progeria Research Foundation\" \/>\n<meta property=\"og:description\" content=\"Lymphoblast Cell Culture Protocols are listed within our website, regarding from shipment of frozen cells and live cultures.\" \/>\n<meta property=\"og:url\" content=\"https:\/\/www.progeriaresearch.org\/ta\/lymphoblast-cell-culture-protocols\/\" \/>\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=\"2025-02-11T14:34:37+00:00\" \/>\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=\"4 minutes\" \/>\n<script type=\"application\/ld+json\" class=\"yoast-schema-graph\">{\"@context\":\"https:\\\/\\\/schema.org\",\"@graph\":[{\"@type\":\"WebPage\",\"@id\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ru\\\/lymphoblast-cell-culture-protocols\\\/\",\"url\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ru\\\/lymphoblast-cell-culture-protocols\\\/\",\"name\":\"Lymphoblast | The Progeria Research Foundation\",\"isPartOf\":{\"@id\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ta\\\/#website\"},\"datePublished\":\"2017-02-27T21:19:33+00:00\",\"dateModified\":\"2025-02-11T14:34:37+00:00\",\"description\":\"Lymphoblast Cell Culture Protocols are listed within our website, regarding from shipment of frozen cells and live cultures.\",\"breadcrumb\":{\"@id\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ru\\\/lymphoblast-cell-culture-protocols\\\/#breadcrumb\"},\"inLanguage\":\"ta-IN\",\"potentialAction\":[{\"@type\":\"ReadAction\",\"target\":[\"https:\\\/\\\/www.progeriaresearch.org\\\/ru\\\/lymphoblast-cell-culture-protocols\\\/\"]}]},{\"@type\":\"BreadcrumbList\",\"@id\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ru\\\/lymphoblast-cell-culture-protocols\\\/#breadcrumb\",\"itemListElement\":[{\"@type\":\"ListItem\",\"position\":1,\"name\":\"Home\",\"item\":\"https:\\\/\\\/www.progeriaresearch.org\\\/\"},{\"@type\":\"ListItem\",\"position\":2,\"name\":\"Lymphoblast Cell Culture Protocols\"}]},{\"@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\":\"ta-IN\"},{\"@type\":\"Organization\",\"@id\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ta\\\/#organization\",\"name\":\"The Progeria Research Foundation\",\"url\":\"https:\\\/\\\/www.progeriaresearch.org\\\/ta\\\/\",\"logo\":{\"@type\":\"ImageObject\",\"inLanguage\":\"ta-IN\",\"@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":"Lymphoblast | The Progeria Research Foundation","description":"Lymphoblast Cell Culture Protocols are listed within our website, regarding from shipment of frozen cells and live cultures.","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\/ta\/lymphoblast-cell-culture-protocols\/","og_locale":"ta_IN","og_type":"article","og_title":"Lymphoblast | The Progeria Research Foundation","og_description":"Lymphoblast Cell Culture Protocols are listed within our website, regarding from shipment of frozen cells and live cultures.","og_url":"https:\/\/www.progeriaresearch.org\/ta\/lymphoblast-cell-culture-protocols\/","og_site_name":"The Progeria Research Foundation","article_publisher":"https:\/\/www.facebook.com\/ProgeriaResearch\/","article_modified_time":"2025-02-11T14:34:37+00:00","twitter_card":"summary_large_image","twitter_site":"@Progeria","twitter_misc":{"Est. reading time":"4 minutes"},"schema":{"@context":"https:\/\/schema.org","@graph":[{"@type":"WebPage","@id":"https:\/\/www.progeriaresearch.org\/ru\/lymphoblast-cell-culture-protocols\/","url":"https:\/\/www.progeriaresearch.org\/ru\/lymphoblast-cell-culture-protocols\/","name":"Lymphoblast | The Progeria Research Foundation","isPartOf":{"@id":"https:\/\/www.progeriaresearch.org\/ta\/#website"},"datePublished":"2017-02-27T21:19:33+00:00","dateModified":"2025-02-11T14:34:37+00:00","description":"Lymphoblast Cell Culture Protocols are listed within our website, regarding from shipment of frozen cells and live cultures.","breadcrumb":{"@id":"https:\/\/www.progeriaresearch.org\/ru\/lymphoblast-cell-culture-protocols\/#breadcrumb"},"inLanguage":"ta-IN","potentialAction":[{"@type":"ReadAction","target":["https:\/\/www.progeriaresearch.org\/ru\/lymphoblast-cell-culture-protocols\/"]}]},{"@type":"BreadcrumbList","@id":"https:\/\/www.progeriaresearch.org\/ru\/lymphoblast-cell-culture-protocols\/#breadcrumb","itemListElement":[{"@type":"ListItem","position":1,"name":"Home","item":"https:\/\/www.progeriaresearch.org\/"},{"@type":"ListItem","position":2,"name":"Lymphoblast Cell Culture Protocols"}]},{"@type":"WebSite","@id":"https:\/\/www.progeriaresearch.org\/ta\/#website","url":"https:\/\/www.progeriaresearch.org\/ta\/","name":"\u0baa\u0bc1\u0bb0\u0bcb\u0b9c\u0bc0\u0bb0\u0bbf\u0baf\u0bbe \u0b86\u0bb0\u0bbe\u0baf\u0bcd\u0b9a\u0bcd\u0b9a\u0bbf \u0b85\u0bb1\u0b95\u0bcd\u0b95\u0b9f\u0bcd\u0b9f\u0bb3\u0bc8","description":"\u0b95\u0bc1\u0bb4\u0ba8\u0bcd\u0ba4\u0bc8\u0b95\u0bb3\u0bc1\u0b95\u0bcd\u0b95\u0bbe\u0b95 \u2665 \u0b9a\u0bbf\u0b95\u0bbf\u0b9a\u0bcd\u0b9a\u0bc8\u0b95\u0bcd\u0b95\u0bbe\u0b95","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":"ta-IN"},{"@type":"Organization","@id":"https:\/\/www.progeriaresearch.org\/ta\/#organization","name":"\u0baa\u0bc1\u0bb0\u0bcb\u0b9c\u0bc0\u0bb0\u0bbf\u0baf\u0bbe \u0b86\u0bb0\u0bbe\u0baf\u0bcd\u0b9a\u0bcd\u0b9a\u0bbf \u0b85\u0bb1\u0b95\u0bcd\u0b95\u0b9f\u0bcd\u0b9f\u0bb3\u0bc8","url":"https:\/\/www.progeriaresearch.org\/ta\/","logo":{"@type":"ImageObject","inLanguage":"ta-IN","@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\/ta\/wp-json\/wp\/v2\/pages\/1004","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.progeriaresearch.org\/ta\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/www.progeriaresearch.org\/ta\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/www.progeriaresearch.org\/ta\/wp-json\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/www.progeriaresearch.org\/ta\/wp-json\/wp\/v2\/comments?post=1004"}],"version-history":[{"count":0,"href":"https:\/\/www.progeriaresearch.org\/ta\/wp-json\/wp\/v2\/pages\/1004\/revisions"}],"wp:attachment":[{"href":"https:\/\/www.progeriaresearch.org\/ta\/wp-json\/wp\/v2\/media?parent=1004"}],"curies":[{"name":"wp (\u0b9f\u0baa\u0bbf\u0bb3\u0bcd\u0baf\u0bc2\u0baa\u0bbf)","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}