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Investigation of nanoscopic dynamics and potentials by interferometric scattering microscopy

표제/저자사항
Investigation of nanoscopic dynamics and potentials by interferometric scattering microscopy / Jaime Ortega Arroyo
Arroyo, Jaime Ortega   
발행사항
Cham, Switzerland : Springer, [2018]
©2018
형태사항
xxxvi, 142 pages : illustrations (some color) ; 25 cm
총서사항
(Springer theses : recognizing outstanding Ph.D. research, ISSN 2190-5053)
주기사항
"Doctoral thesis accepted by the University of Oxford, Oxford, UK"
Includes bibliographical references
표준번호/부호
ISBN 9783319770949
ISBN 3319770942
ISBN 9783319770956 (eBook)
ISBN 3319770950 (eBook)
ISSN 2190-5061 (electronic)
분류기호
듀이십진분류법-> 570.282
주제명
Chemistry    Microscopy    Interferometry
내용유형
text
매체유형
unmediated
수록매체유형
volume

권별정보

권별정보 목록
편/권차 편제 저작자 발행년도 ISBN 청구기호 자료이용하는곳 자료상태
Jaime Ortega Arroyo [2018] 9783319770949 W570.282-19-1 보존 서고 신청후이용(보존)
CONTENTS
1 Introduction = 1
  References = 4
2 Non-fluorescent Single-Molecule Approaches to Optical Microscopy = 7
  2.1 Introduction = 7
  2.2 Single-Particle Tracking = 8
  2.3 Scattering Detection : An Alternative to Fluorescence = 12
  2.4 Interferometric Scattering = 17
    2.4.1 Confocal Detection = 18
    2.4.2 Non-scanned Wide-Field Detection = 19
    2.4.3 Confocal Beam Scanning Wide-Field Detection = 20
  2.5 Applications = 21
    2.5.1 Lateral Single-Particle Tracking = 21
    2.5.2 Axial Localisation via Interferometry = 23
    2.5.3 Label-Free Imaging = 28
  2.6 Conclusion and Outlook = 29
  References = 31
3 Experimental Methods = 37
  3.1 Experimental Optics and Hardware = 37
    3.1.1 Interferometric Scattering Channel = 39
    3.1.2 Focus Control Feedback Channel = 42
    3.1.3 Single-Molecule Fluorescence Channel = 43
    3.1.4 Sample Stage Stabilisation = 43
    3.1.5 Camera Characterisation = 44
    3.1.6 Operation and Synchronisation of the Acousto-Optic Beam Deflector = 46
    3.1.7 Data Acquisition = 47
  3.2 Experimental iSCAT Microscopy = 48
    3.2.1 Image Processing = 48
    3.2.2 Spot Detection = 52
    3.2.3 Localisation = 53
    3.2.4 Trajectory Linking = 54
    3.2.5 Assessment of Localisation Precision = 55
    3.2.6 Self-referencing = 55
  References = 57
4 Anomalous Diffusion Due to Interleaflet Coupling and Molecular Pinning = 59
  4.1 Introduction = 59
  4.2 Experimental Methods = 61
    4.2.1 Materials = 61
    4.2.2 Vesicle Preparation = 62
    4.2.3 Substrate Preparation = 62
    4.2.4 Supported Lipid Bilayer Formation = 62
    4.2.5 Instrument Setup Parameters = 63
  4.3 Experimental Results = 63
    4.3.1 GM1 Undergoes Anomalous Diffusion in Supported Lipid Bilayers = 63
    4.3.2 Transient Confinement Causes Anomalous Diffusion = 66
    4.3.3 Concentration Dependent Dynamics of Transient Binding = 69
    4.3.4 Recovery of Brownian Motion upon Tuning Substrate Interactions and Interleaflet Coupling = 70
  4.4 Discussion = 72
    4.4.1 Importance of Simultaneous Localisation Precision and Time Resolution = 72
    4.4.2 Thermal and Optical Force Considerations = 73
    4.4.3 Membrane Defects, Labelling Artefacts, and CTxB Induced Aggregation Do Not Cause Transient Binding = 74
    4.4.4 Transient Binding Requires Substrate Interaction and Interleaflet Coupling = 75
    4.4.5 Multiple CTxB-GM1 Interactions Result in Ring-Like Structures = 75
    4.4.6 A Model of Transient Binding : Molecular Pinning = 76
  4.5 Conclusion and Outlook = 77
  References = 77
5 Structural Dynamics of Myosin 5a = 81
  5.1 Introduction = 81
  5.2 Experimental Methods = 83
    5.2.1 Sample Preparation = 83
    5.2.2 Experimental Setup= 84
  5.3 Experimental Results = 84
    5.3.1 N-Terminus Labelling Does Not Perturb the Kinetics of Myosin 5a = 84
    5.3.2 During Myosin Movement the Motor Domain Undergoes a Transition Between Two Distinct States = 85
    5.3.3 The Labelled Motor Domain Moves in Three Dimensions = 86
    5.3.4 A Conformational Change in the Motor Domain Accompanies the Power Stroke = 88
    5.3.5 Myosin Steps via a Single, Spatially-Constrained Transient State = 91
    5.3.6 Transient States Occur on the Same Side of Actin for Each Head Domain = 95
    5.3.7 The Diffusion Rate of the Unbound Labelled Head Is Comparable to the Frame Time of 1ms = 96
  5.4 Discussion = 97
    5.4.1 Association Between an N-Terminus Rotation and the Lever Arm Motion = 97
    5.4.2 Sub-steps Along Actin and Leading Head Detachment Do Not Significantly Contribute to the Mechanochemical Cycle = 98
    5.4.3 Myosin Preferentially Walks in a Plane Perpendicular to the Glass Surface = 99
    5.4.4 Structurally Constrained Diffusion Leads to Unidirectional Motion = 101
    5.4.5 Relationship Between the Transient State and the AB Transition = 103
    5.4.6 Directionality of the Symmetric Hand-Over-Hand Mechanism = 104
  5.5 Conclusion and Outlook = 105
  References = 106
6 All Optical Label-Free Detection, Imaging and Tracking of Single Proteins = 111
  6.1 Introduction = 111
  6.2 Experimental Methods = 113
    6.2.1 Experimental Setup Parameters = 113
    6.2.2 Sample Preparation = 113
  6.3 Experimental Results = 113
    6.3.1 Label-Free Detection of Actin Filaments = 113
    6.3.2 Label-Free Detection of Single Proteins = 114
    6.3.3 Detection Sensitivity of iSCAT = 116
    6.3.4 Comparison of Single-Molecule Fluorescence and iSCAT Imaging = 117
    6.3.5 Observation of Single-Molecule ATP-Dependent Kinetics = 119
    6.3.6 Nanometric Tracking of Individual Myosin Molecules = 119
  6.4 Discussion = 120
  6.5 Conclusion and Outlook = 121
  References = 122
7 Single-Molecule Chemical Dynamics : Direct Observation of Physical Autocatalysis = 125
  7.1 Introduction = 125
  7.2 Experimental Methods = 126
    7.2.1 Experimental Setup Parameters = 126
    7.2.2 Sample Preparation = 126
  7.3 Results and Discussion = 127
    7.3.1 Detection of Micellar Aggregates as the Product of the Chemical Reaction = 127
    7.3.2 Super-Resolution Imaging of the Progress of the Reaction = 129
    7.3.3 Direct Observation of Bilayer Formation = 131
    7.3.4 Characterisation of the Reaction Kinetics = 132
    7.3.5 Observation of Physical Autocatalysis in Situ = 133
    7.3.6 Interfacial Dynamics : Surface Interactions Lead to Different Mechanistic Pathways of Product Formation = 136
    7.3.7 Complex Phenomena in the Oil Phase = 137
  7.4 Conclusion and Outlook = 138
  References = 139
8 Outlook = 141
  Reference = 142

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