{"id":17,"date":"2012-06-15T14:48:44","date_gmt":"2012-06-15T13:48:44","guid":{"rendered":"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/?page_id=17"},"modified":"2025-11-24T09:36:28","modified_gmt":"2025-11-24T09:36:28","slug":"protocols","status":"publish","type":"page","link":"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/","title":{"rendered":"Protocols"},"content":{"rendered":"<p>A selection of<em> Trypanosoma brucei<\/em>-specific&nbsp;protocols, updated on an <em>ad hoc<\/em> basis. For details of standard molecular biology protocols see, for example, <em>Current Protocols in Molecular Biology<\/em> or&nbsp;<em>Short Protocols in Molecular Biology, <\/em>both edited by Frederick Ausubel and published by Wiley.<\/p>\n<h2 class=\"wp-block-heading\"><em>Trypanosoma brucei<\/em>&nbsp;culture &amp; manipulation<\/h2>\n<ul class=\"wp-block-list\">\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/bloodstream-form-culture\/\">Bloodstream-form (BSF) culture<\/a><\/li>\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/electroporation\/\">BSF electroporation and selection of transformants<\/a><\/li>\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/bsf-to-procyclic-differentiation\/\">BSF to procyclic differentiation<\/a><\/li>\n<li><a title=\"Procyclic-form culture\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/procyclic-form-culture\/\">Procyclic-form culture<\/a><\/li>\n<\/ul>\n<h2 class=\"wp-block-heading\"><em>Trypanosoma brucei<\/em> growth analysis<\/h2>\n<ul class=\"wp-block-list\">\n<li>Basic growth assay (&amp; calculating a doubling time)<\/li>\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/ec50-analysis\/\">EC<sub>50<\/sub> analysis<\/a> (calculating a &#8216;half maximal effective growth-inhibitory concentration&#8217;)<\/li>\n<li>Isobologram analysis (synergistic, additive &amp; antagonistic inhibitor interactions)<\/li>\n<\/ul>\n<h2 class=\"wp-block-heading\">Molecular biology<\/h2>\n<ul class=\"wp-block-list\">\n<li>&#8216;Sticky end&#8217; cloning (getting a PCR amplified insert into a plasmid)<\/li>\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/phenolchloroformisoamyl-alcohol-extraction-and-ethanol-precipitation\/\">Phenol:chloroform:isoamyl alcohol extraction and ethanol precipitation of DNA<\/a><\/li>\n<li>Electroporating <em>E. coli<\/em> and identifying recombinant&nbsp;clones<\/li>\n<\/ul>\n<h2 class=\"wp-block-heading\">Cell biology<\/h2>\n<ul class=\"wp-block-list\">\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/immunofluorescence\/\">Immunofluorescence<\/a><\/li>\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/mitotracker-staining\/\">Mitotracker staining<\/a> (localising the mitochondrion)<\/li>\n<li>DAPI cell cycle analysis<\/li>\n<li>FACS analysis<\/li>\n<\/ul>\n<h2 class=\"wp-block-heading\">Protein analysis<\/h2>\n<ul class=\"wp-block-list\">\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/protein-preparation\/\">Protein preparation<\/a><\/li>\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/hypotonic-lysis\/\">Hypotonic lysis<\/a>&nbsp;(soluble and &#8216;membrane-associated&#8217; fractions)<\/li>\n<\/ul>\n<h2 class=\"wp-block-heading\">RNA analysis<\/h2>\n<ul class=\"wp-block-list\">\n<li><a title=\"html\" href=\"http:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/page-separation-of-small-rnas\/\">PAGE separation of small RNAs<\/a><\/li>\n<li>Transcriptional run-on analysis (detecting nascent RNAs in <em>Trypanosoma brucei<\/em>)<\/li>\n<\/ul>\n","protected":false},"excerpt":{"rendered":"<p>A selection of Trypanosoma brucei-specific&nbsp;protocols, updated on an ad hoc basis. For details of standard&#8230;<\/p>\n","protected":false},"author":116,"featured_media":0,"parent":0,"menu_order":3,"comment_status":"closed","ping_status":"closed","template":"","meta":{"footnotes":""},"class_list":["post-17","page","type-page","status-publish","hentry","odd"],"yoast_head":"<!-- This site is optimized with the Yoast SEO plugin v26.9 - https:\/\/yoast.com\/product\/yoast-seo-wordpress\/ -->\n<title>Protocols - Alsford Lab<\/title>\n<meta name=\"robots\" content=\"index, follow, max-snippet:-1, max-image-preview:large, max-video-preview:-1\" \/>\n<link rel=\"canonical\" href=\"https:\/\/blogs.lshtm.ac.uk\/alsfordlab\/protocols\/\" \/>\n<meta property=\"og:locale\" content=\"en_GB\" \/>\n<meta property=\"og:type\" content=\"article\" \/>\n<meta property=\"og:title\" content=\"Protocols - Alsford Lab\" \/>\n<meta property=\"og:description\" content=\"A selection of Trypanosoma brucei-specific&nbsp;protocols, updated on an ad hoc basis. 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