title: Interferometric Scattering Microscopy for the Study of Molecular Motors.
abstract: Our understanding of molecular motor function has been greatly improved by the development of imaging modalities, which enable real-time observation of their motion at the single-molecule level. Here, we describe the use of a new method, interferometric scattering microscopy, for the investigation of motor protein dynamics by attaching and tracking the motion of metallic nanoparticle labels as small as 20nm diameter. Using myosin-5, kinesin-1, and dynein as examples, we describe the basic assays, labeling strategies, and principles of data analysis. Our approach is relevant not only for motor protein dynamics but also provides a general tool for single-particle tracking with high spatiotemporal precision, which overcomes the limitations of single-molecule fluorescence methods.
authors: Physical and Theoretical Chemistry Laboratory, University of Oxford, Oxford, United Kingdom.
authors: J
authors: J
authors: Andrecka
authors: Laboratory of Molecular Physiology, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD, United States.
authors: Y
authors: Y
authors: Takagi
authors: Pennsylvania State University, University Park, PA, United States; Intercollege Graduate Degree Program in Bioengineering, Pennsylvania State University, University Park, PA, United States.
authors: K J
authors: KJ
authors: Mickolajczyk
authors: Pennsylvania Muscle Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, United States.
authors: L G
authors: LG
authors: Lippert
authors: Laboratory of Molecular Physiology, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD, United States.
authors: J R
authors: JR
authors: Sellers
authors: Pennsylvania State University, University Park, PA, United States; Intercollege Graduate Degree Program in Bioengineering, Pennsylvania State University, University Park, PA, United States.
authors: W O
authors: WO
authors: Hancock
authors: Pennsylvania Muscle Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, United States.
authors: Y E
authors: YE
authors: Goldman
authors: Physical and Theoretical Chemistry Laboratory, University of Oxford, Oxford, United Kingdom. Electronic address: philipp.kukura@chem.ox.ac.uk.
authors: P
authors: P
authors: Kukura
mesh_terms: D004398:Dyneins / Q000737:chemistry / Q000302:isolation & purification*; D006801:Humans; D016547:Kinesins / Q000737:chemistry / Q000302:isolation & purification*; D008856:Microscopy, Fluorescence / Q000379:methods*; D008857:Microscopy, Interference / Q000379:methods; D020409:Molecular Motor Proteins / Q000737:chemistry / Q000378:metabolism; D009218:Myosins / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: C488101:KIF5B protein, human; D020409:Molecular Motor Proteins; D009218:Myosins; D004398:Dyneins; D016547:Kinesins
keywords: Dynein; High speed; Interferometric scattering microscopy; Kinesin; Molecular motors; Myosin; Single molecule; Single-particle tracking