Microliter to nanoliter scale amino acid assays of eye fluids

The ocular system is nourished and protected by eye fluids, such as tears, aqueous humor and vitreous humor. The chemical composition of those eye fluids, such as protein, peptides, amino acids, ions and other small molecules reflects metabolism and intercellular communication of the ocular tissues, such as cornea and retina. The changes of chemical composition in eye fluids may be induced by the eye diseases and resulted in pathological metabolism and cellular alterations. The goal of this thesis is to develop analytical methods for amino acid analysis of eye fluid samples from microliter to nanoliter range to investigate the changes of amino acid content corresponding to the physiology of ocular tissues and may provide insights into the progression of eye diseases.;Capillary electrophoresis with laser induced fluorescence detection (CE-LIF) is a well-established analytical method. A rapid and reliable separation method is developed for amino acid assays of human vitreous and is used to explore the basic physiology of the retina and for further investigation of the progression in diabetic retinopathy.;An elution method was developed and employed to the Schirmer strip test for tear chemical analysis. This is the first demonstration of CE applied to tear amino acid analysis following a Schirmer strip test. The tear amino acid assay may suggest a new line of research into the very common dry eye syndromes.;A nano-scale manual sample handling method is developed to aliquot rat vitreous perfusate from each rat vitreous sample of low flow push pull perfusion sampling technique for nitrate and amino acid analysis with different detection methods. The rat aqueous humor analysis by an optimized direct sampling method is presented in this thesis as well. Both preliminary results from developed sampling techniques provide exceedingly promising future for developing animal model to understand basic physiology and further applied to investigate the pathogenesis of eye diseases. The methods developed for amino acid assays of eye fluids in this thesis can be widely employed to investigate the physiology of the ocular tissues and to explore the progression of eye diseases in the future.