Instrumentation and techniques in confocal microscopy

Abstract

Liquid crystal variable focal length microlenses are designed, fabricated and tested for inclusion into the confocal system. Their emerging light fields are imaged and examined. This proves a novel, efficient and thorough examination of the optical properties, the results of which are compared to those found using interferometric methods. The aberrations present are noted and possible causes discussed. It is also found that by increasing the driving frequency of the applied voltage across the liquid crystal lenses the optical performance can be enhanced by up to 30%, allowing diffraction-limited microlenses to be achievable at greater focal length ranges. The inclusion of these lenses as a high throughput aperture array with a fibre bundle as the corresponding pinhole array at the detector is experimentally investigated. The use of the microlens arrays as an array of objective lenses is also discussed and the depth discriminating properties examined by use of the axial response. The fibre bundle is shown to be an efficient pinhole array and an imaging conduit in confocal systems. The mapping of the entrance and exit positions of an incoherent fibre bundles is undertaken for the proposed application as a pinhole and image conduit. Using fluorescent confocal and conventional microscopy, defect analysis and the size determination of dye-doped melamine formaldehyde microspheres by spectral analysis is performed. High contrast confocal fluorescent lifetime imaging of gold nanoparticles by novel sample preparation is examined and the possible uses of this technique in the future discussed.