Biochemistry, Biophysics, and Structural Biology Commons^™

Interaction Of Laminin Beta2 With Eps8 And Desmin In Muscle Cells, Luke Xueliang Cui

Theses, Dissertations and Capstones

Laminin β2 is localized to the synaptic basal lamina (BL) in muscle and glomerular BL in kidney. To find interacting proteins, a mouse kidney cDNA library was screened with domain I of rat laminin β2 using the yeast two-hybrid system. Fifteen positive clones were found. The DNA sequence of three each of these clones corresponded to the phosphoprotein Eps8 (epidermal growth factor receptor pathway substrate clone 8, which is tyrosine phosphorylated after EGF stimulation) and the intermediate filament (IF) protein keratin. To determine if this interaction occurred in muscle cells, C2 cells transfected with the rat …

Lateral Hypothalamic Nmda Receptor Subunits Nr2a And/Or Nr2b Mediate Eating: Immunochemical/Behavioral Evidence, Arshad Khan, Margarita Curras, Jennifer Dao, Faizi Jamal, Rishi Goel, Elizabeth Gillard, Stefany Wolfsohn, B. Stanley

Selected Works Temporary Series

No abstract provided.

Archaeal Aminoacyl-Trna Synthesis: Unique Determinants Of A Universal Genetic Code?, Michael Ibba, A. W. Curnow, J. Bono, P. A. Rosa, C. R. Woese, D. Söll

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

No abstract provided.

Lateral Hypothalamic Nmda Receptor Subunits Nr2a And/Or Nr2b Mediate Eating: Immunochemical/Behavioral Evidence, Khan, Margarita Curras, Jennifer Dao, Faizi Jamal, Rishi Goel, Elizabeth Gillard, Stefany Wolfsohn, B. Stanley

Departmental Papers (Biology)

No abstract provided.

The Atomic Force Microscope In The Elucidation Of Leukocyte Abnormality, Derbrenn O'Connor

Theses

The aim of this project was to investigate the novel use of the atomic force microscope (AFM) as a technique in the elucidation of leukocyte abnormality. To this end, comparative AFM imaging and elasticity studies were performed on normal and myeloid leukaemic leukocytes.

Initially, an evaluation of techniques for the isolation of specific leukocyte (e.g. lymphocytes, granulocytes, etc.) populations from whole blood or leukocyte concentrates, was carried out. It was found that a silanised glass surface failed to select and immobilise a leukocyte population. However, centrifugation of whole blood in an Isopaque-ficoll gradient solution yielded a viable mixed leukocyte population …

Plasmd Bearing A Cdna Copy Of The Genome Of Bovine Viral Diarrhea Virus, Chimeric Derivatives Thereof, And Method Of Producing An Infectious Bovine Wral Darrheavirus Using Sad Plasmid, Ruben O. Donis, Ventzislav B. Vassilev

School of Veterinary and Biomedical Sciences: Faculty Publications

A plasmid bearing a cDNA copy of the genome of bovine viral diarrhea virus (BVDV), chimeric derivatives of the plasmid and a method of producing an infectious bovine viral diarrhea virus using the plasmid are disclosed. The invention relates to a plasmid DNA molecule that replicates easily in E. coli and contains a sufficient portion of the genome of BVDV, cloned as cDNA, to be a suitable template to produce RNA in vitro which, upon transfection into bovine cells, gives rise to infectious BVDV. The BVDV created by the process of the invention can be engineered for use as a …

In Situ Hybridization For The Detection And Localization Of Swine Chlamydia Trachomatis, C. Chae, D.-S. Cheon, D. Kwon, O. Kim, B. Kim, J. Suh, D. G. Rogers, K. D. E. Everett, A. A. Anderson

School of Veterinary and Biomedical Sciences: Faculty Publications

Gnotobiotic piglets were inoculated intralaryngeally with swine Chlamydia trachomatis strain R33 or orally with swine C. trachmatis strain R27. Archived formalin-fixed, paraffin-embedded tissues from piglets euthanatized 4–7 days postinoculation were examined by in situ hybridization for C. trachomatis nucleic acid using a nonradioactive digoxigenin-labeled DNA probes that targeted specific ribosomal RNA or omp1 mRNA molecules of the swine C. trachomatis strains. Positive hybridization signals were detected in bronchial epithelial cells, bronchiolar epithelial cells, pneumocytes, alveolar and interstitial macrophages, and jejunal and ileal enterocytes. Chlamydia-infected cells had a strong signal that was confined to the intracytoplasmic inclusions. Positive hybridization signals were …

Affinity Chromatography: A Review Of Clinical Applications, David S. Hage

David Hage Publications

Affinity chromatography is a type of liquid chromatography that makes use of biological-like interactions for the separation and specific analysis of sample components. This review describes the basic principles of affinity chromatography and examines its use in the testing of clinical samples, with an emphasis on HPLCbased methods. Some traditional applications of this approach include the use of boronate, lectin, protein A or protein G, and immunoaffinity supports for the direct quantification of solutes. Newer techniques that use antibody-based columns for on- or off-line sample extraction are examined in detail, as are methods that use affinity chromatography in combination with …

Urinary N-Acetyl-Beta-D-Glucosaminidase In Rheumatoid Arthritis, M P. Iqbal, A A. Ali, M A. Waqar, N Mehboobali

Department of Biological & Biomedical Sciences

Excretion of urinary N-acetyl beta-D-glucosaminidase (NAG) and its isoenzyme patterns were studied in two groups of patients with rheumatoid arthritis (RA) and in normal control subjects. Urine samples were collected from 30 seropositive RA patients, 19 seronegative RA patients, and 15 normal healthy subjects. All the patients and normal subjects were assessed to have normal liver and kidney functions. A small portion of the urine sample was dialyzed against 0.01 M phosphate buffer, pH 7.0 and NAG activity was monitored. Mean +/- SD values of urinary NAG in seropositive RA patients, in seronegative RA patients and in normal healthy subjects …

Localization Of Sodium, Potassium-Adenosine Triphosphatase In The Cerebral Cortex Of Developing Rats, Maki Ujiie

Loma Linda University Electronic Theses, Dissertations & Projects

Active transport via Na⁺, K⁺-ATPase plays an important role in establishing and maintaining proper ionic gradient across all mammalian cell membranes. A strict regulation of Na⁺, and K⁺ ion levels is especially important in excitable cells such as neurons and myocytes. Inhibition of the enzyme is associated with several pathological conditions and has serious detrimental effects on cerebral cortical cells. Yet the enzyme appears to be limited in amount and distribution in newborn rats. In this study, Na_⁺, K⁺- ATPase expression and distribution in early postnatal rats were studied …

In Vitro Assessment Of The Toxicity Of Cocaine And Its Metabolites In The Human Umbilical Artery, Tessa L. Long

Electronic Theses and Dissertations

An in vitro model was used to assess the effect of cocaine and its metabolites on the umbilical artery. Objectives were to pharmacologically confirm the presence of adrenergic innervation using tyramine, evaluate the ability of cocaine, benzoylecgonine, norcocaine and cocaethylene to potentiate vasoconstriction by serotonin and norepinephrine, examine the ability of ketanserin to block the enhanced vasoconstriction produced by cocaine, and determine displacement of 3 H-ketanserin by serotonin, norepinephrine, tyramine and mianserin. The vasoconstrictive effect of tyramine (100 μM) was enhanced in the presence of cocaine by 257%. Vasoconstrictive effects of serotonin and norepinephrine were significantly enhanced by cocaine by …

Expression, Isolation And Purification Of Human Zona Pellucida Protein 3, Ting-Fung Chi

Theses and Dissertations in Biomedical Sciences

Mammalian fertilization involves interactions of sperm surface receptors with the ligands of the zona pellucida, an extracellular matrix surrounding the ovulated oocytes. In humans, the zona pellucida is composed of three major glycoproteins. One of them, ZP3, participates in the primary sperm binding and in the subsequent triggering of the spermatozoa's acrosome reaction. Studies on the role of this specific protein in the human fertilization process are hampered by the limited amount of available biologically functional proteins.

By use of a pcDNA 3.1(+) expression vector, a transfecting-vector was constructed containing a 1.3 kb histidine tagged hZP3 cDNA. This histidine tagged …

Development Of Fluorescent Techniques In The Kinetic Characterization Of Reverse Transcriptase, Kevin Kevin Howerton

Chemistry & Biochemistry Theses & Dissertations

Reverse transcriptase (RT) is an RNA-directed DNA polymerase isolated from human immunodeficiency virus (HIV) and other retroviruses. It has been the primary target for anti-HIV research since the discovery of its vital role in the retroviral life-cycle. The conventional method for carrying out kinetic characterization and inhibition studies of reverse transcriptase involves the use of radioactive materials. Although this method is effective, it is both time consuming, due to a necessary repeated washing step, and hazardous to the worker and environment. Furthermore, strict laws limit the use of radioactive materials to licensed workers and laboratories. In response to these concerns, …

Leukemia Inhibitory Factor (Lif): Murine Preimplantation Embryo Development, Implantation Rates, And Skeletal Development, Michael Hayes Mitchell

Theses and Dissertations in Biomedical Sciences

Leukemia inhibitory factor (LIF) is a pleiotropic cytokine which demonstrates perplexing physiological effects. It has been demonstrated that LIF is essential for implantation in mice. Little is known relating to the manner by which LIF effects pre-implantation and post-implantation development. The objectives of this project were to determine the effects LIF on pre-implantation development, to determine the effects that it may have on implantation rates, successful pregnancy rates, and resorption rates, and to determine the effects that LIF has on the skeletal development of mice. For the embryo transfer experiments, embryos were exposed to test compounds in the transfer medium …

Characterization Of The Biological Functions Of Human Recombinant Zona Pellucida Protein 3, Yu Wen Juan

Theses and Dissertations in Biomedical Sciences

Recombinant human zona pellucida protein 3 (rhZP3), expressed. isolated and purified from PA-1 cells, is characterized for its biological activity and the role in the signal transduction pathway. Characterization of the biological activity of rhZP3 was detected by hemizona assay and immunofluorescence staining of acrosome reaction The results indicated that rhZP3 exhibited an inhibition in the binding assay (HZI 43.6 +/-3.3; n = 9; 30 ng/mL rhZP3) and induction of acrosome reaction (198.6% +/- 77.2% increase from baseline; n = 29; 30 ng/mL rhZP3). It was further confirmed by the transmission electron microscopy that there was no difference in morphology …

A Model Using Radiation And Pws4-Htnf-Α Gene Therapy For Treatment Of Glioblastomas, Angelo G. Baher

Loma Linda University Electronic Theses, Dissertations & Projects

The efficacy of radiotherapy for cancer is limited by the dose that can be safely delivered to the tumor without causing debilitating side effects. In addition, successful treatment of highly malignant tumors such as glioblastomas is likely to require adjunctive therapies to enhance tumor response to radiation. Previous studies have shown immunomodulation and a synergestic reduction in tumor volume of malignant tumors when tumor necrosis factor-α (TNF-α) protein is administered prior to radiation. The major goal of the present investigation was to evaluate the efficacy of pWS4-human TNF-α (pWS4-hTNF-α), a new plasmid construct that expresses human TNF-α protein, together with …

The Second Messenger Camp Elicits Eating By An Anatomically Specific Action In The Perifornical Hypothalamus, Elizabeth Gillard, Arshad Khan, Rickinder Grewal, Bara Mouradi, Stefany Wolfsohn, B. Stanley

Selected Works Temporary Series

No abstract provided.

The Second Messenger Camp Elicits Eating By An Anatomically Specific Action In The Perifornical Hypothalamus, Gillard, Arshad Khan, Rickinder Grewal, Bara Mouradi, Stefany Wolfsohn, B. Stanley

Departmental Papers (Biology)

No abstract provided.

Developmental And Hormonal Regulation Of The Expression Of Cytosolic Phospholipase A2 Protein Levels In Baboon Placenta, Kathleen Elizabeth Dunstan

Chemistry & Biochemistry Theses & Dissertations

Cytosolic phospholipase A₂ ( cPLA₂) specifically catalyzes the release of arachidonate from membrane phospholipids, thus initiating synthesis of prostaglandins, thromboxanes, and other eicosanoids. The human and nonhuman primate placenta actively synthesize high levels of eicosanoids which play both an autocrine and paracrine role in placental function. Previous studies in Dr. Miriam D. Rosenthal's laboratory have shown that cPLA₂ mRNA levels are developmentally regulated. The present study was designed to determine whether placental cPLA₂ protein levels increased during gestation. Since the marked increases in estrogen levels during normal pregnancy have been shown to enhance morphological and …

Enhancement Of Atp Levels And Glucose Metabolism During An Infection By Chlamydia: Nmr Studies Of Living Cells, David M. Ojcius, H. Degani, Joel Mispelter, Alice Dautry-Varsat

All Dugoni School of Dentistry Faculty Articles

The Chlamydia species are obligate intracellular bacteria that proliferate only within the infected cell. Since the extracellular bacteria are metabolically inert and there are no cell-free systems for characterizingChlamydia metabolism, we studied metabolic changes related to ATP synthesis and glycolysis in HeLa cells infected withChlamydia psittaci during the course of the 2-day infection cycle using noninvasive 31P and 13C NMR methods. We find that the infection stimulates ATP synthesis in the infected cell, with a peak of ATP levels occurring midway through the infection cycle, when most of the metabolically active bacteria are proliferating. The infection also stimulates synthesis of …

Land Use Patterns In Relation To Lake Water Quality In The Great Pond Watershed, Problems In Environmental Science Course (Biology 493), Colby College, Colby Environmental Assessment Team, Colby College

Colby College Watershed Study: Great Pond (2012, 2010, 1998)

Lakes are natural resources, which have many effects on the land surrounding them, They support adjacent communities by providing water and regulating temperatures, helping to define the surrounding ecosystem, and serving as sources of drinking water as well as recreation. The prolonged presence of human activity in a watershed can disturb the physical and chemical cycles of the lake and its surrounding ecosystems (Henderson-Sellers and Markland 1987). Over time, lakes undergo a process called eutrophication, a natural aging process during which the nutrient levels increase and dissolved oxygen levels decrease (Smith and Smith 1998). As the lake ages or becomes …

Eating Induced By Perifornical Camp Is Behaviorally Selective And Involves Protein Kinase Activity, Elizabeth Gillard, Arshad Khan, Bara Mouradi, Omkar Nalamwar, B. Stanley

Selected Works Temporary Series

No abstract provided.

Evaluation Of The Role Of Fructosamine As An Indicator Of Diabetic Control, Katherine Hooley

Theses

Diabetes mellitus, a common metabolic disease, is characterised by excess circulating glucose concentrations. Such a hyperglycaemic status provides the stimulus for irreversible glycation (non-enzymatically post-translational) of structural and functional proteins. Protein glycation, although a cause of complication (neuropathy, micro/macrovascular disease), provides a time-averaged index of diabetes status and control. In particular, glycated plasma proteins analysis, i.e., the fructosamine assay, has received favourable response due to its speed, simplicity, low cost, and ease of automation. However, lack of standardisation and a universally-accepted calibrant has limited technical and subsequent clinical confidence in fructosamine analysis. In response, a comprehensive technical evaluation (including novel …

Development Of An Atomic Force Microscopic Immunosensor For The Ferritin Antigen, Brendan O'Connell

Theses

The adsorption conditions for a range of different immunoreactants to both polystyrene and ultrasmooth silanised silicon surfaces are optimised. The conditions studied were concentration of protein, time and temperature of incubation, pH, molarity and ionic strength of the buffer. The anti-ferritin surface was selected as our model system. This immunosensor was used to develop and evaluate a millispot ELISA for ferritin. The millispot ferritin ELISA developed utilises a sample volume of 6pl and a sample incubation time of 10 minutes. Precision and accuracy studies show the assay to have a dynamic assay range of 7.8-lOOng/ml ferritin. The millispot ELISA detected …

Development Of A Differential Assay To Selectively Measure Α-Amylase Isoenzymes., Miriam O'Connor

Theses

α-Amylase is a heterogeneous polypeptide, primarily found in the Salivary and Pancreatic glands, giving rise to two isoenzymes, namely Salivary- and Pancreatic-type Amylases. Total α-Amylase activity measurements are routinely carried out in hospital laboratories to aid in the diagnosis of diseases of the pancreas and in the investigation of pancreatic function in patients with severe abdominal pains. With the development and wide availability of modem chromogenic oligosaccharide substrates specific for Amylase, assays have become more sensitive, rapid and easier to use.

Differential Amylase Isoenzyme assays ware developed to distinguish isoenzyme levels. Several methods were developed for isoenzyme determination. Of these. …

Development Of An Elisa Method For The Measurement Of Cd44 Expression In Human Serum, Ciara Hurley

Theses

CD44 is a cell surface glycoprotein (mol. wt. =85-95 kDa) which has been implicated in lymphocyte-HEV interactions and the process of lymphocyte homing. CD44 is a complex glycoprotein which undergoes extensive alternative splicing to generate various CD44 isoforms. CD44 functions include the delivery of an activation signal to T-cells, binding to hyaluronate, mediation of lymphocyte homing, and the spread of metastatic cells. CD44 has been particularly implicated in the dissemination of non- Hodgkin's lymphoma and as such may serve as a useful prognostic marker of disease progression. CD44 measurement in serum may provide a rapid, inexpensive and relatively non-invasive prognostic …

Development And Validation Of An Improved Ultrasensitive Assay For Cardiac Troponin I (Ctni), William G. Blake

Theses

Cardiac troponin I (cTnl) has been reported to be a highly specific marker for myocardial injury. In patients suffering from acute coronary syndromes, a sensitive serum marker which is closely related to the extent of myocardial damage which provides prognostic data and which can be measured rapidly is desirable. In this thesis a “sandwich” enzyme immunoassay was developed to measure cTnI. The developed assay was reproducible with the inter assay precision ranging from 4.8-14% and the intra assay precision ranging from 3.1-9.5%. The limit of detection of the assay was calculated as 0.385 ng/ml. The upper reference range (non-parametric 95% …

Eating Induced By Perifornical Camp Is Behaviorally Selective And Involves Protein Kinase Activity, Gillard, Arshad Khan, Bara Mouradi, Omkar Nalamwar, B. Stanley

Departmental Papers (Biology)

No abstract provided.

Polyadenylation Of Vesicular Stomatitis Virus Mrna Dictates Efficient Transcription Termination At The Intercistronic Gene Junctions, Leroy N. Hwang, Nathan Englund, Asit K. Pattnaik

School of Veterinary and Biomedical Sciences: Faculty Publications

The intercistronic gene junctions of vesicular stomatitis virus (VSV) contain conserved sequence elements that are important for polyadenylation and transcription termination of upstream transcript as well as reinitiation of transcription of downstream transcript. To examine the role of the putative polyadenylation signal 3' AUACU₇5' at the gene junctions in polyadenylation and transcription termination, we constructed plasmids encoding antigenomic minireplicons containing one or two transcription units. In plasmid-transfected cells, analyses of the bicistronic minireplicon containing the wild-type or mutant intercistronic gene junctions for the ability to direct synthesis of polyadenylated upstream, downstream, and readthrough mRNAs showed that the AUACU …

Development And Validation Of Flow-Injection (Continuous-Flow) Elisa Techniques., Declan Gerard Spillane

Theses

A flow-through enzyme-linked immunosorbent assay was developed based on affinity chromatography using the determination of ferritin in serum as a model system. In this method, samples and standards are introduced to separate columns containing immobilised anti-ferritin antibody, and antigen bound by solid-phase antibody is subsequently detected using an anti-ferritin-alkaline phosphatase conjugate. To detect immobilised label, p-nitrophenyl phosphate is added and product is developed in the column at room temperature. Following elution of product from the column, the absorbance is measured and the columns are regenerated using a low pH elution. The final developed system requires approximately 1.5 h for the …