This thesis looks at fluorescence techniques and their use for nanometrology applications. It has been primarily industrially linked with scientific instrument vendors Horiba Scientific IBH and Horiba Scientific ISA and examines the state of the art instrumentation taking the reader from grounds up from simple steady-state and time-resolved fluorescence spectroscopy techniques that are routinely used and newer more advanced techniques made possible but rapid developments in technology. Secondary industrial links are to NPL allowing the examination of the state of the art instrument in fluorescence microscopy an extension of fluorescence spectroscopy and this thesis likewise builds from simple to advanced microscopy techniques.In the case of nanometrology well-established techniques in particular time-resolved fluorescence anisotropy, which overcomes the diffraction barrier by use of low concentrations and infers the average particle size of a homogeneous distribution by use of polarized light and Brownian motion are discussed. These applications are examined in conjunction to high concentration microscopy techniques such as direct Stochastic Optical Reconstruction Microscopy (dSTORM) High concentration heterogeneous techniques are better suited to most biological applications which involve the measurement of highly packed nanostructures. dSTORM requires use of specific chemical conditions and high laser power to enable stochastic blinking. A video of these stochastic blinks by use of a fast capture imaging CCD allows one to temporally resolve each single-molecule blinks and construct a single super-resolution image. The implication of these chemical conditions, high laser power and limitations even in today’s state of the art CCDs need to be properly understood and any development in either but preferably all three will make this advanced microscopy technique more feasible.This work looks at the properties of some new probes for nanometrology however, the strict criteria required for successful dSTORM applications in particular leaves this work an open investigation.
|Date of Award||8 Apr 2016|
- University Of Strathclyde
|Sponsors||HORIBA Jobin Yvon IBH Ltd|
|Supervisor||David Birch (Supervisor) & Yu Chen (Supervisor)|