Chemical imaging and spectroscopy using tunable filters: Instrumentation, methodology, and multivariate analysis

Spectral imaging has experienced tremendous growth during the past ten years and is rapidly becoming a formidable analytical tool. Recent advances in electronically tunable filters and array detectors are enabling high resolution spectral images to be acquired of chemical and biological systems that have traditionally been difficult to study non-invasively. Additionally, the development of powerful and inexpensive computer platforms is broadening the appeal of spectral imaging methods which have historically required costly and computationally adept computer workstations. The emphasis of my research has been to explore high throughput widefield imaging instrumentation and methodology using novel acousto-optic tunable filter (AOTF) and liquid crystal tunable filter (LCTF) imaging spectrometers. In order to demonstrate the feasibility of employing multiplexed AOTFs for spectroscopy and chemical imaging applications, a near-infrared (NIR) multiplexed AOTF spectrometer employing Hadamard encoding sequences has been developed. In addition, the use of multiplexed AOTFs as adaptive filters in NIR spectroscopy and fluorescence imaging has been demonstrated.; A second type of electronically tunable image filter, the liquid crystal tunable filter (LCTF) has recently been developed and is well suited to high resolution, diffraction limited imaging applications. The earliest generation of LCTFs was based on the Lyot birefringent filter and possessed small transmittances due to the use of multiple polarizers and imperfect waveplate action. An improved LCTF prototype incorporating split-element Lyot filter stages has been evaluated and compared to the earlier generation of LCTF devices.; The high image fidelity, wide acceptance angle, and large clear aperture of the LCTF make it well suited to macroscopic chemical imaging applications. A macroscopic imaging fluorometer employing LCTFs for source tuning and emission filtering has been developed for high throughput microtiter plate based assays. The instrument has been incorporated into a commercial microtiter plate reagent dispenser and can image the fluorescence emission from microtiter plates at rates up to 10 frames/second. The instrument design and its evaluation using model fluorophores is described in detail.; The final emphasis of my research has been to explore and develop rapid multivariate analyses that complement the high throughput acquisition methods employed in our laboratory. A new technique called cosine correlation analysis (CCA) is introduced which rapidly generates image contrast based on spectral shape. The theory and implementation of CCA are described using model data and Raman image data from thermoplastic olefin and silicon semiconductor materials.