General Experimental Methods 2.1 Introduction 2.2.1 Glass Cleaning

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Abstract	The majority of the assays in this thesis involved in vitro reconstitutions with dynamic microtubules and pre-polymerized actin filaments. These were typically immobilized on a glass surface and imaged by total internal reflection fluorescence (TIRF) microscopy. This chapters outlines the general methods applicable to all the in vitro assays described in this thesis. 2.2 Flow cell preparation and surface functionaliza-tion To minimize non-specific binding of fluorescent proteins to the glass, hence to avoid too much background signal, we made use of thoroughly cleaned glass slides and coverslips, which were then functionalized through a variety of techniques. Glass coverslips were cleaned in base piranha solution, a safer alternative to acid piranha solution that only becomes reactive when heated at or above 60 • C. Storing the cleaned coverslips in a 0.1 M KOH aqueous solution activated hydroxyl (OH −) groups on the 35
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