Anti Gfap Synthesis

The research into Alzheimer's Disease has come a long way since 1906 when it is was discovered by Alois Alzheimer. He detected microscopic brain tissue changes called senile and neuritic plaques in deceased patients. These are chemical deposits consisting of protein molecules called Amyloid Precursor Protein(APP) that are fundamental components of a normal brain. However in the brain of an Alzheimer patient, an enzyme cuts the APP apart and leaves fragments in the brain tissue. These combined with degenerating nerve cells cause the plaques or lesions. These lesions are found in many sections of the brain including the hippocampus which regulates emotion and memory, the basal forebrain, and especially the basal nucleus of Meynert and the cortex, where the memory function is located.(2) Another sign of a diseased brain are neurofibrillary tangles, which are malformations within nerve cells.

Based on the samples the expression of rGFP was the greatest in the E3 both quantitatively and qualitatively. The purity of rGFP was however low. The rGFP protein was purified using the Ni+2 agarose chromatography. This part of the experiment did not achieve a high purity. Causes of this may have incorrect pipetting mechanism or incorrect amount deposition. Improving pipetting skills might have lead to more accurate results. A better comprehension of the purpose of each step in the protocol may have resulted in improved results. Also personally having to use GCE of a partner’s could have altered the results. Being more strict with the lab procedures, and not messing up with the timings of certain steps by accident would have improved data. Although minor errors may have occurred the greatest fluorescence occurred in wash fraction W4 and elution fraction E3 quantitatively, and this data matched the qualitative observation of the fluorescence in both these samples. W4 had the greatest protein amount. W1 was the void volume and only contained breaking buffer without rGFP therefore there should have been no fluorescence, and this occurred correctly. Samples W2 and W3 had slight fluorescence which could be explained by the rGFP lacking the His6 tag required to bind to the Ni+2 agarose, or all the binding sites in the column could

the degeneration of brain tissue in mammals. A form of proteins that have undergone alteration form

He observed tangles within the neurons and dense deposits/plaques around the neurons. These deposits and tangles are considered as the two most important pathological hallmarks of the disease.

It has been recently reported that rTMS induces transcription of the glial fibrillary acidic protein (GFAP) in the murine brain. GFAP transcription is up-regulated in astrocytes of the dentate gyrus, and the magnitude of the response depends on the number of stimulus trains (5). Whether rTMS induces GFAP transcription in astrocytes directly or indirectly through neural activation remains to be determined.

Alzheimer's is a neurodegenerative disorder that affects the neurons within the brain, causing memory loss as well as language, and behavioral disturbances. An Alzheimer's brain contains Amyloid plaques that form toxic proteins which are called amyloid beta. The amyloid plaques start to spread throughout the brain causing tangles to form in the brain. The tangles are a form of twisted threads of protein. In compartment to a “normal” brain with an Alzheimer's brain, that Alzheimer's brain is remarkably smaller in size and contains fluid flied spaces.

Alzheimer 's disease is a severe illness that affects the brain and leads to gradual memory loss, reduced intellectual ability and deterioration function of thinking. Alzheimer 's disease (AD), the most common type of dementia, is a progressive neurological disorder that increasingly robs individuals of cognitive, behavioral, and functional skills (Demakis, 2007). The reason for the appearance this disease is a collection of disorders in the brain due to which its cells are dying partially. In the case of the early stage of Alzheimer disease when the disease is not carried out the appropriate treatment mental functions are destroyed completely. The disease is diagnosed mainly in elderly people. The symptoms of Alzheimer 's disease, were first isolated by German psychiatrist Alois Alzheimer, is one of the most common forms of dementia. “As is well known, Alzheimer first described the disease that would eventually be named for him at a meeting of the South West German Psychiatrists in Tubingen in 1906. It was a brief report of the case of a 51year old woman who developed progressive dementia, accompanied by focal signs, hallucinations and delusions. On post-mortem, her brain was found to contain numerous senile plaques and a newly observed pathological structure – densely twisted bundles of neurofibrils, or neurofibrillary tangles, which were made visible to microscopic observation through a newly developed silver-staining technique.” (Perry, G. et. all, 2006). Typically, the

To obtain the cheek cell a piece had to be scraped off the interior of a person mouth with a toothpick. Next, using a pipette, a drop of water was placed onto the slide. Then after, the side of the toothpick, which was inside of the person mouth was placed onto the water on the slide. Also to for the cheek cell to be seen a yellowish iodine was placed into the water on the slide. Next a coverslip was placed on the edge of the slide then lowered over the sample and , was carefully lowered into place using tweezers. This method help prevent air bubbles from being trapped under the coverslip. Now that the slide was prepared the observation could begin. The slide was placed onto the stage of the microscope. To begin the observation the microscope was placed at low power. At a low power (40x) you could see the individual cells, such as the cell membrane, cytoplasm, and the skin of the cell. Next, the microscope was switched to a medium power (100x), through the microscope everything was clearer, the nucleus was visible with a couple other cells being able to be seen . Finally, under high power (400x), things such as a the cytoplasm, cell membrane, and nucleus were seen but on a larger

An initial set of tests is used to differentiate between transudate and exudate fluid; they check fluid protein, albumin, lactate dehydrogenase level, cell count and appearance

The processing of APP involves three proteases, α-, β-, and γ-secretases and two processing pathways, the amyloidogenic pathway and the anti-amyloidogenic pathway as shown in Figure 2 (14, 23). The Aβ is generated in amyloidogenic pathway which is mediated by β-, and γ-secretases (14). The β- secretase first sheds the ectodomain of APP and generates the APP carboxy-terminal fragment (βCTF) (14). Then the γ-secretase cuts at the transmembrane region of βCTF and releases the Aβ into the extracellular fluids like cerebrospinal fluid or plasma (14, 24). In the anti-amyloidogenic pathway α-secretase first cuts at the middle region of Aβ so a trauncated APP CTF (αCTF) is generated and after the transmembrane cut of γ-secretase, a truncated Aβ peptide is released (p3) (Figure 2; 25). The amyloidogenic and anti-amyloidogenic competes with each other (26-27). In addition, when the γ-secretase cuts at the transmembrane region of APP, the cut site is not restricted to a single position. The γ-secretase first cuts at

Fluorescence spectroscopy is a useful technique to determine the proteins and nucleic acids of a macromolecule. Some molecules, fluorophores, can absorb light at certain wavelengths and then emit it at another wavelength. When the molecule absorbs light it is excitation and the light released is emission. The light particles, photons, hit a fluorophore to excite the molecule (Voet et. al 2013). Excitation occurs by the valence electrons of the molecule receiving energy and moving to a higher energy state from the ground state. The excited electrons will later return to the ground state because they

Microscopy is the most extensively used imaging technique to discover biological systems, in which immunocytochemistry (ICC), the localization of proteins inside cells, and immunohistochemistry (IHC), which discloses the distribution, profusion, and localization of biomarkers within a tissue, can be observed directly. Immunohistochemistry (IHC) mentions to the process of detecting antigens in cells of a tissue section by exploiting the principle of antibodies binding specifically to antigens in biological tissues With IHC, tissues are removed from the patient either frozen or chemically well-preserved and fixed in paraffin. Segments as thin as 4μm are sliced from frozen or paraffin-embedded tissue and fixed onto slides in

In the central nervous system, microglia and astrocytes are the major type of glial cells, and activation of these cells has been involved in all neurodegenerative diseases (Wyss-Coray and Mucke, 2002). Nevertheless, the diverse physiological functions of glial activation might complicate the interpretation of experimental

We used Orco-Gal4 and 10x; UAS-CD8; GFP to drive GFP into the ORNs. We used primary antibody that was a chicken anti-GFP (1:150, Invitrogen) and rabbit anti GABA (B) R1 (sequence mentioned above) in 0.2%PPST+5% normal goat serum (Triton X-100). Samples were incubated overnight at 4 degrees C. Secondary antibodies were a goat anti-chicken Alexa 488 (1:150, Invitrogen) and a goat anti-rabbit Alexa 647 (1:75, Invitrogen) in 0.2% PBST+5% normal goat serum. GFP staining was clearly observed in ORN’s which is represented in Figure 10. This result suggests that ORN’s project much deeper into the brain than thought previously. Next, we looked for the presence of GABA (B) receptors in the larval antennal lobe. Immunohistochemistry analysis were performed in animals expressing GFP in all 21 ORNs. We found that GABA (B) receptor R1 localized to the ORN synaptic terminals. We conclude that the GABA (B) R1 is localized at the ORN synapses. Root et al., 2008 observed GABA (B) receptors on ORNs in adult flies. Here with our results, consistent with Root et al, we are seeing GABA (B) receptors staining in larval stage too. Staining image is represented in Figure 11.

Methods used in this experiment include use of plasmids, imaging techniques, use of HEK cells, and voltage/current clamps, and finally an SNRC calculation. Plasmids are used because they are easy to manipulate and integrate into plasma membranes1. This allows an efficient and quick insertion of desired substances into the desired protein like GFP into ASAP1. In this experiment GFP was introduced into the S3 S4 loop where