A series of sulfonate anionic lipids esters derived from 4-sulfobenzoic acid (single chain) or 5-sulfoisophthalic acid (double chain) with chain length from C14 to C18 were synthesized and characterized. The sodium salts were uniformly insoluble in ethanol; the tetramethylammonium salts of the single chain derivative from oleyl alcohol and the double chain derivative from 2-octyldodecan-1-ol were sufficiently soluble for subsequent experiments. Lipids in ethanol and aqueous buffers were mixed in a microfluidic system (NanoAssmblr ® microfluidic mixer) to prepare a lipid dispersion containing vesicles and/or nanoparticles. Initial studies on prediction and controlling vesicle size based on lipid geometric parameters showed that particle size could be successfully affected and controlled by altering lipid compositions consistent with the formation of vesicles. A survey using high resolution cryo-Scanning Transmission Electron microscopy of the sample made by the microfluidic mixer demonstrated that vesicles were formed but a majority of the sample reformed to other aggregates, which complicated the interpretation of the initial product distribution. Further investigation on the efficiency of incorporation of phospholipids into vesicles indicated that 55% of the initial phospholipid appeared in the vesicle fractions. Sulfonate anionic lipids are incorporated into vesicles with lower efficiency and reach a threshold beyond which the sulfonate lipid is not incorporated. Entrapment efficiency was studied with three dyes. Different concentrations of the hydrophobic neutral dye Nile red, the hydrophilic cationic dye neutral red and the hydrophilic anionic dye hydroxypyrene trisulfonate (HPTS) were prepared. The entrapment efficiency was quantitatively analyzed by HPLC, and electrospray mass spectrometry; up to 15% of the initial dye present could be entrapped. Vesicles permeability assays using the ion channel gramicidin and the ion carrier valinomycin with HPTS-loaded vesicle samples showed that vesicle samples made by the microfluidic mixer and made by a conventional extrusion method appeared to behave in the same manner. Addition of a sulfonate anionic lipid to the lipid mixture resulted in vesicle leakage. The unilamellar proportion of HPTS loaded vesicle samples was assessed using a mellitin assay. A vesicle sample made by the microfluidic mixer was 80% unilamellar; a vesicle sample made by the extrusion method on the same lipid mixture was 60% unilamellar. / Graduate / mengxiuzheng@gmail.com

published_or_final_version / Anatomy / Master / Master of Philosophy

No description available.

Access to abstract permanently restricted to Ball State community only / Access to thesis permanently restricted to Ball State community only / Department of Physiology and Health Science

Vesicle associated membrane protein 2 (VAMP 2) is a synaptic vesicle protein involved with exocytosis in many different cell types, such as pancreatic cells, parotid salivary cells, adrenal cells, skeletal cells, and adipocytes. Also, white blood cells such as eosinophils, neutrophils, and mast cells have been characterized to process VAMP 2. In this study, we tested the hypothesis that VAMP 2 is associated with the vesicle population in rabbits B lymphocytes and may serve as the v-SNARE for vesicular antibody release. Two Rabbit B lymphoblast cell lines were used to detect the presence of VAMP 2, which are the 240 E IgG secreting plasmacytoma-like cell line and 55D1 IgM surface expressing cells. The cell lines were broken down into vesicle and plasma membrane fractions. Immune dot blots demonstrated VAMP 2 was positive in the vesicle fraction of both cell lines. However, VAMP 2 was expressed more by the 240 E IgG secreting cell line. Western blots displayed diverse results with bands that ran at or below 20 KDa, which is consistent with the known molecular weight bands for VAMP 2 of 12.6 kDa and 18 kDa. Our results suggested that VAMP 2 is associated with the vesicle population in rabbit B lymphocytes and could serve as the v- SNARE for vesicular antibody release. / Access to thesis permanently restricted to Ball State community. / Department of Physiology and Health Science

VAMP-7 has been found to interact with SNAP-23, a t-SNARE that functions in relocating granule membranes in response to stimulation, and plays a large role in the regulation of granule release from mast cells in response to an allergic reaction. While evidence suggests that VAMP-7 is active in antibody release in the innate immune system, little investigation has been completed on VAMP-7 interaction in specific antibody release of B lymphocytes of the humoral immune system. Little research has previously focused on vesicular transport within B lymphocytes, leaving molecular mechanisms within B lymphocytes a mystery. Immunodot blots, western blots, and immunoflourescent microscopy were all utilized with the goal of identifying the presence of VAMP- 7. Immunobot blots for both 55D1 and 240E cells were all negative for the presence of VAMP-7. However, VAMP-7 was detected using immunoflourescent microscopy in both 55D1 and 240E cell lines. / Access to thesis permanently restricted to Ball State community. / Department of Physiology and Health Science

Thesis (Ph.D.). / Written for the Dept. of Anatomy and Cell Biology. Title from title page of PDF (viewed 2008/05/08). Includes bibliographical references.

Thesis (Ph. D.)--University of Texas at Austin, 2005. / Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Washington, 2000. / Vita. Includes bibliographical references (leaves 122-130).

Thesis (Ph. D.)--Cornell University, May, 2008. / Vita. Includes bibliographical references.