Matching Items (37)

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Sequence-specific detection of different strains of LCMV in a single sample using tentacle probes

Description

Background
Virus infections often result in quasispecies of viral strains that can have dramatic impacts on disease outcomes. However, sequencing of viruses to determine strain composition is time consuming and

Background
Virus infections often result in quasispecies of viral strains that can have dramatic impacts on disease outcomes. However, sequencing of viruses to determine strain composition is time consuming and often cost-prohibitive. Rapid, cost-effective methods are needed for accurate measurement of virus diversity to understand virus evolution and can be useful for experimental systems.
Methods
We have developed a novel molecular method for sequence-specific detection of RNA virus genetic variants called Tentacle Probes. The probes are modified molecular beacons that have dramatically improved false positive rates and specificity in routine qPCR. To validate this approach, we have designed Tentacle Probes for two different strains of Lymphocytic Choriomeningitis Virus (LCMV) that differ by only 3 nucleotide substitutions, the parental Armstrong and the more virulent Clone-13 strain. One of these mutations is a missense mutation in the receptor protein GP1 that leads to the Armstrong strain to cause an acute infection and Clone-13 to cause a chronic infection instead. The probes were designed using thermodynamic calculations for hybridization between target or non-target sequences and the probe.
Results
Using this approach, we were able to distinguish these two strains of LCMV individually by a single nucleotide mutation. The assay showed high reproducibility among different concentrations of viral cDNA, as well as high specificity and sensitivity, especially for the Clone-13 Tentacle Probe. Furthermore, in virus mixing experiments we were able to detect less than 10% of Clone-13 cDNA diluted in Armstrong cDNA.
Conclusions
Thus, we have developed a fast, cost-effective approach for identifying Clone-13 strain in a mix of other LCMV strains.

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Agent

Created

Date Created
  • 2017-10-13

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Diurnal Cycle Modeling of Nutrient Transport through the Intervertebral Disc to Prevent Future Degeneration after Transplantation

Description

The intervertebral disc goes through degenerative changes with age, which leads to disc thinning, bulging, or herniation. Spinal fusion treatments are ineffective as they cause quicker degeneration of adjacent discs

The intervertebral disc goes through degenerative changes with age, which leads to disc thinning, bulging, or herniation. Spinal fusion treatments are ineffective as they cause quicker degeneration of adjacent discs and fail in nearly 20% of cases, so researchers have turned to tissue-engineering biocompatible intervertebral discs for transplantation. However novel and effective as this may seem, these transplanted discs still show evidence of degeneration after just 5 years. I hypothesize that these discs are degenerating due to a blockage of the cartilaginous endplates post-transplantation that is hindering nutrient transport through the intervertebral disc. In order to test this hypothesis, I developed a mathematical model of nutrient transport through the intervertebral disc in one diurnal daily loading cycle. This model was used to simulate open endplates and blocked endplates and then compare differences in nutrient concentration and nutrient transport to the center of the disc. Results from the math model simulations were then compared to in vitro experimental data collected in lab to verify the findings on a physiological level. Results showed significant differences, both in vitro and in the model, between nutrient transport in open endplates vs blocked endplates, lending support to the original hypothesis. This study only presents preliminary results, but could hold the key to preventing future disc degeneration post-transplantation.

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Created

Date Created
  • 2015-05

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Insulin Resistance in Rats Exposed to a High Fat Diet

Description

Type II diabetes is a serious, chronic metabolic disease that has serious impacts on both the health and quality of life in patients diagnosed with the disease. Type II diabetes

Type II diabetes is a serious, chronic metabolic disease that has serious impacts on both the health and quality of life in patients diagnosed with the disease. Type II diabetes is also a very prevalent disease both in the United States and around the world. There is still a lot that is unknown about Type II diabetes, and this study will aim to answer some of these questions. The question posed in this study is whether insulin resistance changes as a function of time after the start of a high fat diet. We hypothesized that peripheral insulin resistance would be observed in animals placed on a high fat diet; and peripheral insulin resistance would have a positive correlation with time. In order to test the hypotheses, four Sprague-Dawley male rats were placed on a high fat diet for 8 weeks, during which time they were subjected to three intraperitonal insulin tolerance tests ((NovoLogTM 1 U/kg). These three tests were conducted at baseline (week 1), week 4, and week 8 of the high fat diet. The test consisted of serially determining plasma glucose levels via a pin prick methodology, and exposing a droplet of blood to the test strip of a glucometer (ACCUCHEKTM, Roche Diagnostics). Two plasma glucose baselines were taken, and then every 15 minutes following insulin injection for one hour. Glucose disposal rates were then calculated by simply dividing the glucose levels at each time point by the baseline value, and multiplying by 100. Area under the curve data was calculated via definite integral. The area under the curve data was then subjected to a single analysis of variance (ANOVA), with a statistical significance threshold of p<0.05. The results of the study did not indicate the development of peripheral insulin resistance in the animals placed on a high fat diet. Insulin-mediated glucose disposal was about 50% at 30 minutes in all four animals, during all three testing periods. Furthermore, the ANOVA resulted in p=0.92, meaning that the data was not statistically significant. In conclusion, peripheral insulin resistance was not observed in the animals, meaning no determination could be made on the relation between time and insulin resistance.

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Created

Date Created
  • 2017-05

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Investigation of Student Achievement and Attitude about a Flipped Classroom Using Linked Lecture Videos in Biomedical Engineering

Description

Flipped classrooms invert the traditional teaching methods and deliver the lecture online outside of the classroom. An increase in technology accessibility is increasing the prevalence of this teaching technique in

Flipped classrooms invert the traditional teaching methods and deliver the lecture online outside of the classroom. An increase in technology accessibility is increasing the prevalence of this teaching technique in universities. In this study, we aim to address some of the uncertainties of a flipped classroom by implementing a new lecture format in Transport Phenomena. Transport Phenomena is a junior level biomedical engineering course originally flipped in Spring 2013. Since transitioning to a flipped classroom, students have been required to watch 75-minute lectures outside of class where the instructor covered key concepts and examples using paper and marker on a document camera. In class, students then worked in groups to solve problems with instructor and teaching assistant feedback. Students also completed self-graded homework with the opportunity to earn lost points back by discussing fundamental misconceptions. We are introducing re-formatted mini lectures that contain the same content broken down as well as example problems worked out in a tutorial technique instead of traditional solving method. The purpose of this study is to determine the effectiveness of newly created mini lectures with integrated questions and links in terms of student achievement and attitude [interest, utility, and "cost" (time, effort, and emotion)].

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Agent

Created

Date Created
  • 2016-12

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Blood Flow as a Biomarker for Diet Induced Thermogenesis

Description

Adaptive thermogenesis is an innate mechanism that assists the body in controlling its core temperature that can be stimulated in two ways: cold and diet. When adaptive thermogenesis is stimulated

Adaptive thermogenesis is an innate mechanism that assists the body in controlling its core temperature that can be stimulated in two ways: cold and diet. When adaptive thermogenesis is stimulated through diet, the metabolic rate of the body should increase and the metabolic efficiency of the body should decrease. This activation should, theoretically, help to control weight gain. A protocol was developed to study four male Sprague-Dawley rats throughout a fourteen week period through the measurement of brown adipose tissue blood flow and brown adipose tissue, back, and abdomen temperatures to determine if diet induced thermogenesis existed and could be activated through norepinephrine. The sedative used to obtain blood flow measurements, ketamine, was discovered to induce a thermal response prior to the norepinephrine injection by mimicking the norepinephrine response in the sympathetic nervous system. This discovery altered the original protocol to exclude an injection of norepinephrine, as this injection would have no further thermal effect. It was found that ketamine sedation excited diet induced thermogenesis in periods of youth, low fat diet, and early high fat diet. The thermogenic capacity was found to be at a peak of 2.1 degrees Celsius during this time period. The data also suggested that the activation of diet induced thermogenesis decreased as the period of high fat diet increased, and by week 4 of the high fat diet, almost all evidence of diet induced thermogenesis was suppressed. This indicated that diet induced thermogenesis is time and diet dependent. Further investigation will need to be made to determine if prolonged high fat diet or age suppress diet induced thermogenesis.

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Created

Date Created
  • 2016-12

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New Diagnostic Methods for Detecting Microvillus Inclusion Disease

Description

Microvillus Inclusion disease is a fatal disease found in the Navajo population caused by a single nucleotide polymorphism. It is characterized by intractable diarrhea and is often fatal early in

Microvillus Inclusion disease is a fatal disease found in the Navajo population caused by a single nucleotide polymorphism. It is characterized by intractable diarrhea and is often fatal early in life.1 The current method of diagnosis is sending duodenal biopsies for histopathological examination and confirmatory testing through genomic sequencing. The purpose of this experiment was to create a more simple and cost-effective diagnostic method for detecting Microvillus Inclusion disease. Three methods were explored (RFLP2, ARMS3,4, and Tentacle Probes5,6) and two methods were tested to determine their ability and their efficiency in detecting the SNP that causes the disease.2 Tests using the RFLP2 method and synthetic DNA resulted in 9% false-positive rate and 11% false-negative rate in a blind trial for detecting both target (mutation present) and non-target (mutation absent) DNA when gel analyzing software was used to compare Rf values after gel electrophoresis. Using the ARMS method3, a nine-sample randomized test was run that ended up with 22% false-positive rate and 19% false-negative rate from a blind trial when using a gel analyzing software to determine presence of the SNP by band intensity. Disclaimer: No DNA from human patients was used in this study. Only synthetic DNA used.

Contributors

Agent

Created

Date Created
  • 2018-05

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Sickle Cell Disease and the Need for Neonatal Testing Programs in Africa, with an Emphasis on Kenya

Description

Sickle cell disease is a genetic disorder that can cause substantial helath problems. It is the result of a mutation in the DNA coding for hemoglobin. As a result of

Sickle cell disease is a genetic disorder that can cause substantial helath problems. It is the result of a mutation in the DNA coding for hemoglobin. As a result of changes in two important amino acids, a person suffering from sickle cell disease will have erythrocytes that do not maintain the typical biconcave shape and instead for a crescent shape. Individuals with sickle cell disease may have many health problems tied to their irregular hemoglobin. The unusual shape of the erythrocytes leads to a much shorter cell life, which means that even though bone marrow remains active long past childhood to try to keep up with the loss of erythrocytes, the body is still unable to accommodate the rapid death of erythrocytes. The malformed erythrocytes can also cause vascular occlusion, blocking blood vessels and slowing blood flow. While sickle cell disease has the potential to spread worldwide, it is particularly common in Africa. This may be because people with the sickle cell trait have a high resistance to malaria, making them more likely to survive that ubiquitous disease and pass on their traits to their offspring. However, the mortality rate in young children with sickle cell disease is very high, in part because the spleen, already stressed by filtering out dead erythrocytes, has difficulties filtering out bacteria. One of the keys to stopping the spread of the disease is neonatal screening, but this requires specialized equipment that is fairly uncommon in rural areas, as can be seen in Kenya. Therefore, it would be highly beneficial to develop a more cost-effective and widely available method for testing for sickle cell disease.

Contributors

Agent

Created

Date Created
  • 2012-05

Imaging Local Drug Delivery

Description

Imaging analysis of local drug delivery is important because in both studies involving chemotherapy targeted toward glioblastoma and antimicrobial addressing infection, the drug concentration and distribution are unknown. There are

Imaging analysis of local drug delivery is important because in both studies involving chemotherapy targeted toward glioblastoma and antimicrobial addressing infection, the drug concentration and distribution are unknown. There are a variety of studies focused on the local delivery of drug to a targeted location, but we are presenting a way of quantifying the concentration of the drug and the distribution of the drug during a period of time. This study aims to do that by utilizing Materialise Mimics to analyze the MRI images of local drug delivery in glioblastoma in canines and antimicrobial gel in rabbit femurs. The focus of the technique is to register the anatomy in T1-weighted spin echo images to the drug delivery in T2 flow attenuated inversion recovery (FLAIR) images in order to see where the drug went and did not go relative to the anatomical part. Both studies focus on addressing effective volumes of drug to a designated anatomical area, in which the delivery can be difficult as it involves bypassing the blood brain barrier in the first study and achieving effective volumes while preventing toxicity to the kidneys in the second study. The goal of this project lies in determining the drug volumes and location for the specified duration and anatomical part.

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Agent

Created

Date Created
  • 2018-05

Improving Medical Aerosols: An exploration of nebulous discharge

Description

Improving medical aerosols is the multifaceted objective that is the overarching theme of this work. This thesis is the culmination of many hours of academic research. It details the current

Improving medical aerosols is the multifaceted objective that is the overarching theme of this work. This thesis is the culmination of many hours of academic research. It details the current mechanical and physiological obstacles of state of the art drug inhalation technology, as well as provides a detailed guide of the experimental set up, procedure, analysis and background for the charge neutralization experiments performed by the author. The findings of this research are that inhalation devices need to become personalized; meaning adjustable flow rates, particle sizes, and charge levels. To improve the efficiency of lung deposition they could use MRI to take advantage of 3D modeling software to make transport models of an individual patient's lungs. This model would allow an engineer to calculate the air velocity in each passage of the respiratory system and would account for any pulmonary obstructions that would completely alter the deposition pattern from the average healthy patient. With the velocity profile of the lung a doctor could formulate an aerosol with the perfect attributed for the most targeted delivery. For the experiments performed in this work the following results were obtained. The ionization of air by polonium 210 alpha particles is dependent on the distance from the alpha emitting source and the strength of the electric field. Furthermore discharge of aerosol droplets is possible through volume conduction however the mass of the polonium 210 isotope must be proportional to the ionization current demand.

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Agent

Created

Date Created
  • 2014-05

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Sickle Cell Disease Education and Screening in Kenya

Description

Sickle Cell Disease (SCD) is a prevalent genetic disease in Africa, and specifically in Kenya. The lack of available relevant disease education and screening mean that most don't understand the

Sickle Cell Disease (SCD) is a prevalent genetic disease in Africa, and specifically in Kenya. The lack of available relevant disease education and screening mean that most don't understand the importance of getting testing and many children die before they can get prophylactic care. This project was designed to address the lack of knowledge with supplemental educational materials to be partnered with an engineering capstone project that provides a low cost diagnostic test.

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Created

Date Created
  • 2014-05