Acrylamide Coated GlassWe use this protocol is apply a coating of a sterically repulsive, non-interacting acrylamide polymer brush onto microscope glass in order to passivate the glass against protein adhesion.
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PEG Coated GlassWe use this protocol is apply a coating of a sterically repulsive, non-interacting acrylamide polymer brush onto microscope glass in order to passivate the glass against protein adhesion.
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Flat Oil-Water Interfaces
Use this protocol to create a 2D large-area but micron-thin layer of (fluorinated) oil in a ~100um thick microscope chamber. This surface is perfect for performing experiments at surfactant stabilized liquid-liquid interfaces. We use it for a 2D active nematic liquid crystal composed of cytoskeletal proteins.
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Passive Tracer ParticlesPassivate micron sized tracer particles (microspheres) with a poly-Lysine PEG brush. The coating works on 3um and 200nm microspheres and uses economical and commercially available reagents. The spheres have been used in microtubule based experiments in Dogic Lab and have been found to not interact (stick) with the protein filaments.
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Tubulin Protein From Brain PurificationThis protocol is adapted from the 2003 Popov Tubulin Purification protocol which purifies tubulin using a high molarity PIPES buffer. Here, I have adapted this protocol for Dogic Lab into step-by-step directions along with notes at each step from one of the early preps.
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Tubulin Protein Recycling GuideOnce tubulin protein is purified, it should be recycled so that only polymerization-competent tubulin remains in the final stock before use in experiments. This protocol is adapted from the widely used Tim Mitchison Lab tubulin and microtubule protocols as a step-by-step guide for the Dogic Lab.
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Tubulin Protein Labeling Guide To use tubulin or microtubules in fluorescence microscopy experiments, I use this protocol to label the tubulin with Alexa 647 dye. The protocol is adapted from the widely used Tim Mitchison Lab tubulin and microtubule protocols as a step-by-step buide for labeling tubulin in the Dogic Lab. This protocol is for a large amount of tubulin and requires a significant amount of dye. It can be scaled to smaller preps easily.
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