To Pledge or not to Pledge Research Paper Example | Topics and Well Written Essays - 1500 words

To Pledge or not to Pledge - Research Paper Example According to Nelson (2012), the enduring significance of the Nightingale Pledge may be attributed to the legacy of the first nursing icon, Florence Nightingale, in her unwavering dedication to cater the needs of the sick and needy, particularly during difficult times, such as war, famine, disease, and natural calamities (pp. 10-11). Considering the eminent influence of the Nightingale Pledge on a nurse’s life, this paper attempts to uncover the underlying historical significance of the said pledge. Further, this paper also attempts to study the various contexts of the Pledge, especially its functions, purpose, ethical significance, limitations, and criticisms. Historical Role As a profession, nursing is deeply rooted in the society due to its outward protection of the rights of humans, particularly in their rights to life and well-being. In fact, Fagermoen (2005) states that the philosophical foundations of nursing are based on the principles of Humanism wherein nurses, even i n their early not standardized practice, aim to protect, serve, and preserve human life (p. 157). Florence Nightingale embodies those humanistic values in her devotion to tend the wounded soldiers of the Crimean War, and, while alleviating the physical conditions of the soldiers, she also attempts to advance the care systems and psychosocial environment of hospitals so as to make those more conducive to a faster healing process (Fagermoen, 2005, p. 157). During her tenure as a military nurse, she wrote a series of nursing books and notes outlining the basic responsibilities and duties of a nurse, which, in later years, became the theoretical and practical foundations of professional nursing (Kim, 2005, p. 1). The life and works of Florence Nightingale gave birth to the nursing profession, a profession that overcomes barriers of time, place, culture, and religion. For instance, although her works influenced the nursing profession, Florence Nightingale was not the one who wrote the Ni ghtingale Pledge; instead, the pledge was authored by Lystra Gretter, an American nurse (â€Å"The Nightingale Pledge,† n.d.). The pledge was first professed by the 1893 nursing graduates of Detroit’s Harper’s Hospital. Nightingale’s legacy even reached in Japan during the outbreak of the war in 1945 wherein a head nurse in Hiroshima Army Red Cross initiated the recitation of the Nightingale Pledge to restore calm and order in the panic-stricken hospital due to the defeat and surrender of the Japanese militia. Nursing stories around the world relate to the Nightingale Pledge wherein most, if not all, nurses find self-satisfaction in their work through saving the lives of others. Function and Purpose In analyzing the Nightingale Pledge, one can infer that the pledge functions as an ethical guide for nurses in their professional practice while it also aims to indicate the roles, obligations, and limitations of being a nurse. For instance, although the Night ingale Pledge undergoes several legislative revisions and amendments due to socio-cultural concerns, the revised and amended versions of the pledge still maintains the universal nursing standard of conduct, which includes professionalism (â€Å"The Code of Ethics,† 2010, p. xiv). For instance, some of the revisions include the ANA 1950 Code and the 1976 Code; each of which outlines the recommended nurse-patient relationship, which is primarily on a professional level. For instance, the prevailing idea of the revised editions delves on a

Managing Organizational Change Term Paper Example | Topics and Well Written Essays - 2500 words

Managing Organizational Change - Term Paper Example It also offers a basis of evaluating the role of leadership and other change agent in the chance process (Kezar, 2001). An organization in the Financial/Banking Industry derives benefits due to roles played by people in the organization and their attitudes to the change process, especially in condition where there are cases of resistance. Nevertheless, this paper will focus on exploring the organizational change in the Financial/Banking Industry, through a description of management theory and system practices used in the organizations. Organization change is defines as a form of evolution that takes place though a life cycle, without which, the organization would not be able to develop or grow properly. Therefore, the growth calls for an organization to undergo a substantial change in different points of a development process. Influence of change is substantial to employees, organization and transformation, which is at the purpose of both employees and organization like a decision to terminate a section of business. However, the decision is at the best interests of the organization in their long-term objective, but consequences of that closure can have an adverse impact on employees involved, since they will be faced with problems of unemployment. In fact, the impact of the changes on the organization and employees, and the possible alternatives of dealing with the changes is a substantial issue that needs addressing. Background Organization change requires a change agent who refers to person that perpetrates the act or develops a process through which things in the organization become different (Shankar, 2013). For instance, a change occurs in situation, which members of an organization adapt or fail to adapt to dynamic environment, which is considered as a way of sustaining their competitive advantage through flexibility (Shankar, 2013). The role of being a change agent can be undertaken by a HR manager in an organization by implementing organizational change , through a demonstration unique versatility and wide range of skills (Stagl, 2011). The HR manager fostering change in the organization focuses on implementation or execution of activities that deal with employees’ behavior and attitude (Stagl, 2011). Therefore, as a change agent he or she focuses on identifying cues that impede change and the step that can be taken to eliminate these obstacles. There are other benefits that involve ability to forecast and identify need for change and this serves as a significant contributor that enables the organization to succeed and survive in adverse business environment (Kezar, 2001). Research techniques in the evaluation process offer a basis of applying literature critiquing teleological model, which are dominant model within the literature; in fact, they are evaluated in terms of their relevance for given forms of changes, which are proven by numerous studies (Shani, Pasmore & Woodman, 2011). Research techniques in the process of eva luating organization change offers recommendations on ways of challenging potential barriers to change, thereby setting principles that can be applied by leaders to establish a new

John Adams Mini Series Essay Example for Free

John Adams Mini Series Essay The epic HBO miniseries John Adams is far more than a simple historical drama or biographical account of the life of one of the United States’ founding fathers. The series is a glimpse of a period of American history of which many Americans today are unfortunately ignorant. When most modern Americans think of the period of the Revolutionary War and the birth of the country, barring those that study history during school or as an occupation, they think merely of stilted figures that are more myth than reality. When they think of George Washington, they think of a stoic, almost superhuman figure, that is best known for being so honest that he could not tell a lie as a child when he chopped down a cherry tree. When they think of Thomas Jefferson, they think of a genius that so poetically created the country’s most important documents and made the Louisiana Purchase that doubled the country’s size. When common Americans think of Benjamin Franklin, they think that he invented electricity. And, when they think of John Adams, most Americans unfortunately do not really know what to think, and more than anything else, the miniseries sets forth to enlighten the masses. John Adams, while a seminal figure in the birth of the U. S. , is largely unknown by so many of young and old citizens that can rattle off facts of popular culture faster than American history. Like many of the other founding fathers, he is viewed less as human and more as a mythological figure in history, though his actual contributions are not as pronounced as Jefferson, Washington, and even Franklin. The miniseries sheds light on the mystery of John Adams by showing him as he was: a man. Not only do they portray him as a man, they portray him as flawed, awkward at times, easy to anger and to feel slighted, but always honest and upright. He is also ceaselessly prudent and refuses to shy away from work or duty. If anything, this is the main attribute that allowed a hard-working self-made man like John Adams to elevate himself to the level of president, despite all the odds that seemed to be stacked against him. He lacked the charisma of his cousin, Sam, the finances of many founding fathers like Jefferson, and the leadership skills of Washington, but he still managed to elevate himself through hard work and his sense of duty. By portraying John Adams as a regular man that elevated himself through reason and duty, the makers of the miniseries are trying to remove the myth from the founding fathers and replace it with something far more valuable: the truth. Adams, like Jefferson, Washington, and Franklin were merely men at a pivotal time in history with strong ideals and the courage to not back down from them. Adams was also portrayed as having many of the problems and home life of common Americans today. Though his children were on the whole upstanding, he still experienced problems with his son Charles, who drank and caroused and felt neglected by his father. This aspect of Adams’ duty was portrayed excellently in the miniseries, as his wife Abigail and his children often felt the pain of having their father and husband away so often. Adams, too, was not devoid of feeling the absence, and his love for his wife often kept him grounded, if not feeling lonely on the long journeys to Europe. The relationship between Adams and his wife is one of the most humanizing aspects of the film, and it portrays the intimate life of a founding father without any romantic idealization. The Adamses loved each other deeply, and Abigail would often soothe her husband’s doubts and insecurities, while also instilling in him the confidence to succeed. As a strong, intelligent woman in a time where women were expected to live nothing more than a domestic life, the miniseries portrays Abigail as contributing a great deal to her husband’s views, and therefore a great deal in his actions concerning the founding of the country. At times, she would take a hard line with him, though never browbeating him, and she would remind him of his abilities when he would sometimes forget. This helps make modern viewers understand the humanity of life in America over two-hundred years ago, with a reality previously unseen in historical films and television shows. There are also many intricacies of Revolutionary society which are rarely portrayed, but dealt with so well in the miniseries. Fashion is a huge issue, and the wigs that men wore make a modern person wonder about such trends, possibly even questioning current fashion trends. Today, successful men wear suits with ties around their necks, and rarely wear hats. While in Adams’ time, no men went out without their hat, and few men went out without their wigs. While this could be a substitute for male pattern baldness, it seems more like a status symbol, where men can be judged by the elaborate nature of their wigs and their curls. The time spent on such things like the fashion of the French court, the powdering of the wigs, and the white wigs worn by the King of England and those in court seem to suggest an intricacy that most modern Americans may find strange, but can be no stranger than the Windsor knot. This, like many other things in the miniseries help modern viewers understand that while things like fashion changes throughout time, men do not. The miniseries John Adams is about much more than a man or a myth, but it serves as an adept chronicling of history that personalizes events significant to the founding of America. Not only does it humanize John Adams, it helps explain to modern Americans that might not know the contributions he made and how much everything they take for granted is owed to his hard work and sacrifices. The ideals that Adams held sacred are still echoed in the fabric of America, even if Americans cannot see it. The hard work, the individualism, the sometimes inexcusable lack of diplomacy in exchange for brutal honesty are all characteristics that have been respected and emulated by Americans throughout the past two centuries. Adams had few pretensions, and though he had more than a few character flaws, he was human. More than anything else, the miniseries adeptly portrays that history is not merely words in a history book or romanticized in film, but history is the collected actions of humans throughout time, warts and all. History is created by men that are flawed, all the problems and benefits of family and love are the same, and sometimes, a few regular men rise up and change the world, like John Adams.

Effect of Oxidative Stress on HAGE Expression

Effect of Oxidative Stress on HAGE Expression Cancer is a group of diseases which involves uncontrolled cell growth, metastasis and invasion of cells to other tissues via lymph node or blood. Cancer is caused by many factors such as (i) Chemicals such as tobacco, alcohol, asbestos etc., (ii) Viral infections such as Human Papiloma Virus, Epstin Barr Virus, hepatitis B, C etc., (iii) Ionizing radiations such as X-Rays, UV-Rays, Gamma Rays etc., (iv) Hereditary factors such as mutations in BRCA 1 and BRCA 2, p53 mutations, APC gene mutations, Retinoblastoma gene mutation etc., (v) Hormones which promote growth proliferation such as growth hormones, estrogen, progesterone etc. Cancer is classified into malignant and benign tumours. Malignant tumours are mostly liquid tumours which metastasize to other tissues via lymph node and blood. Benign tumours are mostly solid tumours which stay at one particular tissue and seldom migrate. Malignant and Benign tumours are re classified on the type of cell and tissue from which they originate. They are classified as follows: i) Carcinomas: These are the malignancies of the epithelial cells, which line the internal organs of the body and also present on the surface the body such as skin. ii) Sarcomas: These are solid tumours present in connective tissues such as bones and lymph systems (Crowley 2006) Furthermore the tumours are classified according to (i) site of origin as in lung cancer, breast cancer etc., (ii) type of cell such as rhabdomyosarcoma and acute lymphocytic leukaemia. (Ruddon 2007) The progression from a normal cell to malignant one involves the dysregulation of genes implicated in the control of normal proliferation / death over many years. These are normal functions regulated by proto-oncogenes and tumour suppressor genes. Proto-Oncogene And Tumour Suppressor Gene Protooncogene is found to be normally expressed in all vertebrates including humans. It is also found to be expressed in some insects and yeasts. Protooncogenes play an important role regulating normal cell growth and differentiation. They also perform functions such as signal transduction and mitogenic signal execution RAS, MYC, TRK, ERK are some examples of protooncogenes (Novakofski 1991). Oncogenes are derived from protooncogenes during the process of carcinogenesis. The formation of active oncogene leads to abnormal cell proliferation and thus contributes to the formation of tumour (Alitalo and Schwab 1986). A protooncogene can be converted into an oncogene in a variety of ways such as (i)Transduction induced by retroviruses which cause the integration viral DNA with the host. This DNA on translation give rise to viral proteins which cause the activation of protooncogene, thereby leading to cancer. (ii) Mutation within the protooncogene leading to an increased activity of the pr otein. (iii) chromosomal translocation where the gene for proto oncogene is translocated to other loci, which cause its abnormal expression. For eg in the translocation of abl in chromosome 9 to bcr region chromosome 22 as found in Philadelphia chromosome (Rowley 1973) (iv) Amplification induced by gene duplication, mis-regulation of gene which leads to overexpression of protooncogenes. (Todd and Munger 1999) Unlike oncogenes, the tumour suppressor genes normally inhibit cell growth in variety of ways such as (i) repressing the genes important for cell cycle progression. For eg., pRb (retinoblastoma ) controls the G1 stage by being hypophosphoryated and not allowing the release of transcription factor E2F which promotes G1-S transition. (ii) Stopping cell cycle on detecting DNA damage. DNA damage induces p53 which now leaves from its interacting partner Mdm2 and increases the activity of p21. This p21 protein then inactivates cyclin dependent kinases which are essential for cell cycle progression, (iii) Apoptosis may also be promoted by p53 when damage is irreversible. (iv) inducing DNA repair proteins to repair DNA damage and prevent abnormal cell proliferation. (Yoshida, et al. 2000) There are a number of mechanisms by which a cancer cell survives in a body. This is illustrated in the diagram below.(Weinberg and Hanahan, 2000) Treatment The choice of therapy depends upon the location and stage / grade of the tumour, as well as the health of the patient. There are a number of therapies for treating cancer such as surgery, chemotherapy, radiaotherapy, hormonal therapy etc., The deregulation of many genes in cancer cells leads to the over-expression of altered proteins which can be used as biomarker which can provide help for the diagnosis and / prognosis of the treatment but can also be used as potential target for immunotherapy. Cancer antigens and their classification Tumour antigen is an antigenic substance produced in the tumour cells and triggers an immune reaction in the host. They are useful in identifying tumour cells and are used in cancer therapy. Tumour antigens are classified as Cancer testis antigen, Differentiation antigens, Tumour specific unique antigens, Overexpressed Self Antigens, Viral antigens, Post-translationally, Oncofoetal antigens, Idiotypic Antigens as shown in the table below Copied from (Li, et al. 2005) Amongst all these categories, cancer testis antigen represent the most promising group of antigen to be used in future immunotherapeutic interventions due to their restricted expression to mainly to tumour cells with the exception of placenta and testis which are immune-privileged sites and therefore would not pose any real issue for the risk of autoimmunity.(Simpson, et al. 2005) HAGE HAGE also known as DDX43 and CT13 antigen belongs to this category of antigens and was first identified by Martelange et al using cDNA subtraction approach of a human sarcoma cell line.(Martelange, et al. 2000)Using rational hybrid analysis, it was found that HAGE gene is located on chromosome 6. HAGE was found to be overexpressed in several tumours. There is a low expression of HAGE in normal tissues since the expression is regulated by hypermethylation HAGE is also over expressed in tissues such as testis, placenta, and ovaries. HAGE is usually expressed at the m-RNA level though recent studies in melanoma cells suggest that HAGE could also be expressed at the protein level.(Mathieu, et al. 2010) The name DDX43 comes from the fact that HAGE also belongs to another group of protein known as Dead box proteins and referred to as DDX43. (Abdelhaleem 2004) Dead box proteins These proteins were discovered in the late 1980s. (Gorbalenya, et al. 1989)Dead box proteins belong to the family of RNA helicases and play an important role in the transcription, post transcriptional modifications such as splicing, transport, translation, decay and biogenesis of ribosomes. NTP hydrolysis provides energy to these enzymes and they unwind dsRNA or disrupt RNA-Protein interaction. DDX and DHX are widely studied Dead Box Proteins. There are many classes of DDX and HAGE belongs to DDX43. They are so named because they contain the amino acids D-E-A-D (Asp-Glu-Ala-Asp) in one of their motifs namely Motif II. Besides they also contain motifs such as motif I, Q-motif, motif VI, motif Ia, Ib, III, IV and V as shown in the diagram below. These motifs bestow properties such as ATP binding and hydrolysis, RNA interaction, remodelling activity etc to these proteins. The consensus sequence of DEAD box family is shown below Copied from (Linder 2006) Regulation and Function of HAGE Both function and regulation of HAGE have not been studied in great detail (Scanlan, Simpson and Old 2004). In normal tissues HAGE expression is regulated by DNA hypermethylation, Histone modifications such as histone aceylation, histone deacylation and histone methylation. However in cancer, there could be many mechanisms of upregulation of HAGE. The most commonly thought mechanism is the demethylation of DNA which leads to HAGE over-expression. (Roman-Gomez, et al. 2007) HAGE could also be induced due to transformation of normal cells during cancer or due to the action of oncogenes. HAGE might also be induced randomly. HAGE may play a role in pre-mRNA splicing, ribosome biogenesis, transcription and initiation of translation. (Rocak and Linder 2004) This project will focus on Head and Neck Cancer. Head and Neck Cancer Head and Neck cancer are the cancers that arise from the upper aerodigestive tract such as nasal cavity, oral cavity, pharynx, larynx and paranasal sinuses. Most of them are of squamous cell carcinoma type of the head and neck and have been given the acronym -SCCHN or HNSCC. (Argiris, et al. 2008). The diagram of the anatomy of the head and neck is illustrated below Copied from (Wu, et al. 2009) More than half a million patients are diagnosed with head and neck cancer every year. Head and Neck cancer in fact accounts for more than 3-10% of the cancers(Gourin and McMains 2005). Smoking, alcohol, chewing of betel leaf with tobacco and areca are the major risk factors for this cancer. Recently, HPV type 16 has been identified as one of the causes for head and neck cancer and account for 40% (Goodger and McGurk 2000) The progression of this tumour is complicated. It progresses from the normal features to hyperplasia, mild dysplasia, moderate dysplasia, severe dysplasia, carcinoma, invasive carcinoma and metastasis. The progression may be caused by due to genetic instabilities such as loss of heterozygosity of chromosome 9p21, inactivation of of p16 and loss of 3p, loss of 18q, inactivation of PTEN, some translocations, 17Beta heterozygosity or TP53 mutation. (Argiris, et al. 2008) Copied from (Argiris, et al. 2008) Interestingly HAGE was found the John van Geest group to be overexpressed in 40% of HNSCC cancers (unpublished data) and although much of its function is now emerging no information exist at the moment regarding the regulation of its expression. However it has been observed that cells left for longer period of time in the incubator had a significantly higher expression of HAGE at the mRNA and protein level. Aim of this project The observation that HAGE was over-expressed in 40% of HNSCC led to hypothesis that cells under stress i.e lack of nutrients, decrease oxygen level, as well as other factors influence HAGE expression. Moreover it has been found that HAGE is also expressed in 20% of Acute Myeloid Leukemia and 50% Chronic Myeloid Leukemia as well as many solid tumours such as HNSCC (Adams, et al. 2002). This suggests that there must be something in common between these 2 very different forms of cancer. CML and solid tumours are known to have a high expression of reactive oxygen species (ROS), therefore it might be possible that ROS induces HAGE expression. Thus this project will focus on the effect of oxidative stress has on HAGE expression. The effect of other stress such as temperature may also be investigated. METHODS Cell Culture The PCI 13 and PCI 30 cell lines would be supplied by Prof E Tatour. These cell lines are specific for Head and Neck cancer. The cells would be grown in RPMI 1640 +10%(v/v) FCS+2mM L-glutamine and incubated at 37oC and 5%C02 atmosphere. Extraction of m-RNA The culture media would be removed and washed with DPBS(Dulbeccos phosphate buffer saline). Then DPBS will be removed followed by the addition of RNA-STAT 60. RNA pellets would be then retrieved and re-suspended in ddH2O. RNA extracted is then quantified using NANODROP 8000 UV spectrophotometer. RNA would be then added in measurement wells of the spectrophotometer. The spectrophotometer would be read at 260 and 280nm and the amount of RNA will be calculated. The RNA concentration is calculated in  µg/ µl before being adjusted to 1 µg/ µl by ddH2O. RNA samples would be then stored at -80oC and used for conducting RT-PCR and RTq-PCR in the future. RT-PCR (Reverse Transcriptase PCR) From the extracted RNA cDNA will be synthesized. 2  µg of RNA sample will be mixed with 1  µl of oligo-dT primers and diluted to 15  µl of ddH20 in an Eppendorf tube. All RNA samples would follow the same process. UNO Thermoblock will be used to heat the tubes to 70 °C for 5 min which allows the primer-RNA annealing. 10  µl of reverse transcriptase mix, which will be prepared by mixing Muloney Murine Leukemia Virus 5-X buffer, dNTPs, RNasin RNAse inhibitor, M-MLV-reverse transcriptase and ddH20. The tubes will be then pre-heated in a water bath at 39.2 °C for 80 min which allows cDNA synthesis. Tubes will be removed and kept again in the UNO-Thermoblock and heated to 95 °C for five minutes which stops the reaction. These tubes would be stored at -20 °C for future use. Real Time qPCR cDNA generated from RT PCR will be used as a template and the reagents used will be (i) 6.25 µl iQ Sybr Green, (ii) 0.5 µl gene specific sense primer (iii) 0.5 µl gene specific anti-sense primer, (iv) 4.75  µl ddH20 and 0.5 µl cDNA template. These reagent mixtures will be added to all tubes for a specific gene. Samples will be usually carried out in duplicate with a negative control that contains the reaction mixture without cDNA. Rotogene 6000 real-time qPCR analyser will be used to carry out RT qPCR. Relative gene expression using 2ΔCT method will be calculated with the help of the expression of housekeeping genes HPRT-1, HSP-27 and HSP-90. The primers which will be used in real time qPCR experiment is given below. These primers are supplied by mwg-Eurofins. The primers have a stock concentration of 100pmol/ µl and will be diluted to a working concentration of 10pmol/ µl. From this working 0.5 µl (5pM) of primers will be used. Total Protein extraction Cells will be grown to 75% confluence in T75 flasks. Cells will be trypsinised, washed and re-suspended in DPBS and these cells will be then counted using trypan blue. Cells will be diluted to 1X106/ml using DPBS and 5 X106 cells would be taken in a 1.5ml eppendorf. Centrifugation will be done to pellet the cells. The supernatant will be discarded and lysis buffer containing a cocktail of RIPA buffer and 10% protease inhibitor will be added to the pellet. The eppendorf tube will be then placed on a tube rotator at 4 °C for 30 min. It will be then kept on ice for 30 min. Cells will be again centrifuged at 14000 RPM for 30 min at 4 °C. Aliquot of the supernatant will be prepared to be stored at -20 °C for future use. Total Protein Assay Total protein assay will be carried out using BioRad Dc protein assay reagents. Series of dilutions (0.2,0.4,0.5,0.8,1.0,1.5,2.0mg/ml) will be created using stock BSA solution with the concentration of 10 mg/ml. Protein extracts and standards would be tested in triplicates and duplicates respectively. Assays will be performed in 96-well round bottom plates. 25 µl and 200  µl of reagent A and B will be added to all the samples and the samples will be incubated at room temperature for an hour. Proteins will be then read at 750nm and their concentrations would be calculated based on the standard values. Western Blotting 30  µl of sample will be loaded into the wells of SDS PAGE gels. 1X tris-glycine-SDS will be used as the running buffer. A known molecular weight ladder will also be run alongside the samples. Initially 70V current will be applied to aid the migration of proteins through the 5% stacking gel. Once the proteins reach the 10% separating gel, 90V current will be applied. After the proteins run through the gel, they will be transferred to the PVDF membrane with the help of liquid transfer. This membrane will be prepared by washing with 10% methanol for 5 seconds, ddH2O for 5 min and transfer buffer for 10 min. Liquid transfer will be carried out using the following steps. 1. Cold transfer buffer will be used to immerse a gel frame. To this gel frame a sponge pre soaked in transfer buffer will be placed Then a filter card, gel, PVDF membrane, another filter card and a second pre soaked buffer sponge will be placed in order starting from the first filter card. Proteins will be transferred with the help of electric current of 100V applied for one hour. After completion, membranes will be cut and treated with different antibodies.10% (w/v) Marvel milk solution will be used to wash the membrane for one hour at room temperature. This washing blocks the non-specific binding sites. The blocking solution will be then discarded and the antibody diluted to 10% (w/v) in Marvel milk solution will be added. This antibody coated membrane will be agitated overnight on a plate rocker at 4 °C. The next day there would be 3, 10 min washes with TSBT(Tris-Buffered Saline-Tween-20). During these washes the membrane vessel will be agitated in plate orbital shaker at room temperature. After completing the washing process, a secondary antibody specific to the primary antibody will be added. The steps for adding the secondary antibody are same as the primary antibody addition except for the fact that no overnight incubation is required and the secondary antibody will be incubated only for one hour while being agitated on the shaker at room temperature. The marker will also be stained using streptavidin-HRP secondary antibody. After performing the above step the membranes will be developed. Membrane development will be performed by placing the membrane in a tray and washing it with ECL reagent. The membrane would be then exposed for a certain period of time using CCD camera. Assay for ROS-DCFDA Stress Test Stock Solution Preparation The stock solution of Hanks Buffered Salt Solution (HBSS)will be prepared according to the manufacturers protocol. DCFDA(2,7-dchlorofluoroscein -diacetate) Assay To test ROS levels DCFDA test will be performed. Before conducting the DCFDA assay, the standardization of optimum concentrations of H2O2 and DCFDA for detecting cell stress should be performed. It is important to note that H2O2 should be added only sub-lethally and should not be added in proportion which may cause cell death. The PCI 13 and 30 cells will be plated out in two 24 well plates. 1ml of each type of cells plus RPM1640 10% FCS media will be added to 8 wells per plate and will be incubated overnight at 37 °C. The media will be removed the next day and increasing concentration of DCFDA will be added to both plates and the cells will be incubated with Tinfoil wrapping to prevent light exposure and incubated at 37oC for 30 min. The cells will be then washed and then cold HBSS will be added. After this step the cells will be stressed with increasing concentration of H2O2 (see diagram below). The cells will then be again wrapped in tin foil and placed on a rocking platform for 15 min. The same method will be followed for treating other wells with different concentrations of DCFDA. Experiments will be performed at least twice for each cell line. H2O2 will be removed after 15 min and cells will be trypsinised with 100microL trypsin and Versene. 800 microL of DPBS will be added to one of the 2 wells an d will be pipetted thoroughly so as to remove the cells from the well surface. This will be then transferred to the other well and then 1ml of solution would be transferred to Flow associated cell cytometry (FACS) tube for analysis. The same procedure will be carried for other sets of wells. DCFDA fluorescence would be measured using Gallios flow cytometer and results would be analysed in Kaluza program. Time Line For The Project During the month of May, all techniques such as Western Blotting, mRNA/protein extraction, Real Time PCR will be learnt. This time is indicated in red colour in the graph below. This will overlap with the time duration, from May to the end of June, during which hydrogen peroxide experiment will be performed as per written in the method section and the expression of HAGE will be monitored at both the mRNA and protein level. Also from the start of June to the end of July repeat experiments will be performed and if time permits, the effect of thermal stress on HAGE expression will also be investigated. This is indicated in the graph below. SUMMARY Cancer Testis Antigens (CTA) are antigens that are expressed in a variety of tumours. They are usually absent in normal tissues with the exception of testis and placenta where they are expressed as self tolerant antigens. Since these antigens have strong immunogenicity and their expression is mostly restricted to tumours, they are ideal targets for cancer immunotherapy. Therefore much research is ongoing for the identification of CTA. Recently helicase antigen HAGE was identified as a CTA and was found to be over-expressed in HSNSCC, AML and CML. Thus there might be a common link between these different types of cancers. Moreover ROS is known to be induced in all these tumours and it may be possible that ROS causes increased expression of HAGE. Thus the aim of this project would be to investigate the effect of oxidative stress on HAGE expression. HAGE expression would be analysed both at the m-RNA and protein level in two cell lines namely PCI 13 and PCI 30 using techniques such as m-RNA extraction, RT PCR, Western Blotting and Protein assay. The ROS level would be evaluated using the DCFDA assay. Since the mechanism of HAGE regulation is unknown, positive result in the project would help in elucidating a mechanism by which HAGE could be regulated.

Government Intervention in the Memphis City School System Essay

Government Intervention in the Memphis City School System There are many factors that contribute to how teachers will teach their students. The student’s background knowledge, learning style, and interest levels are all very important to take into consideration when constructing a lesson. But teachers have to answer to parents, principles, administrators, school boards, state standards committees, and governmental agencies in addition to their students when it comes fulfilling their job description. One of the most well know government intervention plans that teachers have to adapt to is No Child Left Behind, implemented by the Bush administration in 2002 to help close the gap between under-performing and exemplary schools. The over arching idea is commendable; provide children with a quality education regardless of where they go to school. Schools have to make the grade in order to receive additional government funding and those that consistently fail to hit its goal will completely overhauled in order to achieve improvement. One school system that has been a consistent poor performer is the Memphis City School System. In their desperation to close the achievement gap the Memphis City School Board has surrendered its charter and proposed a vote to merge with its successful counterpart, the Shelby County school system. The surrender of the charter puts the responsibility of providing education for the districts previously run by the city on the shoulders of the Shelby County government, because according to Tennessee state law 49-1-102: â€Å"There shall be a local public school system operated in each county or combination of counties. There may be a local public school system operated in a municipality or special school distri... ...How do you get the community at large to get behind the merge? What will happen to any funds left over from the Gates Foundation donation to the city? Will the consolidation of systems affect the amount of Federal funding provided to the new system through Race to the Top? Personally I think this is a short sighted solution to a problem city school administration and School Board Members cannot seem to find a solution to, which is breaking through the poverty and racial barriers to AYP achievement. Yes, the demographic is unfortunate but forcing this solution on the city and county will not help race relations it will only increase the tension. This merger is another poorly planned government intervention that is bound to fail unless it gains community support and implements a carefully constructed plan for success. We’ll just have to wait and hope for the best.

Ethical Norms of Medical Research Essay -- Pharmaceutical Industry, Br

Branding, according to Parry, is to create a new thinking of the product to customers in the unique way so that the product can compete to others. And, in the pharmaceutical industry, branding is not only creating the new thinking of product, but also defining the best treatment for particular condition. If done so, the condition branding has numerous benefits because it creates consensus internally and externally. Internal consensus which is the activities focus on the problem and solution between brand managers and clinic so that the product can be maximized investment. Beside that, external consensus creates the better relationship between the owner and the clinical community to approach therapeutics (1). However, Fishman argued that branding leads to good sales and profits because the clinical trial researchers are position themselves as mediator between producers, pharmaceutical companies, and consumer, that are clinicians and patients, and they act like experts in that field. Therefore, the researchers create faithfulness to consumer, and in turn branding increase sales (12). As a mediator, researchers involve in the benefit of branding. Somehow, there is a conflict in their role between their benefit and the ethical norm of medical research. Fishman may be right when she implied that the ethical norm of medical research is driven by marketable drugs and marketable diagnoses. Let take a look in the process of how the drugs develop and bring to the market. First of all, the drug which is developed should meet the condition that can treat for disease with certain amount of patients, and then academic medical researchers are person who collect the clinical data, will mediate with FDA to get approval guideline for new drugs; ... ...cluded that With the increasing public†private arrangements being brokered between industry and academia, these ethical issues only become more entangled. Conventional medical ethics is unequipped to deal with these questions. Business ethics is likewise unable to accommodate the particular responsibilities of the biomedical project. The chasm left in between is where the academic researcher now treads, guided only by professional conventions and a growing entrepreneurialism and commercialism in medicine.(24). It may be strict if we say that the drugs market now needs to be control seriously and let the scientist be independent with their research because when drugs consider as other commodity, then patient become as customer not a patient. The patients need to be treated with medical ethical norm so that they can feel they are in treatment with good condition.

Apush Chapter Notes

Although they shared similarities, the Northern and Southern colonies in the 17th and 18th centuries also had many differences. The diversity of the United States goes back to its beginning as a collection of northern and southern colonies. Their differences in religion, politics, economics, and social issues, and the way they dealt with them, are what shaped our country into what we are today. Religion in the southern colonies was not practiced with the enthusiasm that it was in New England. While most colonists of the south were Anglicans, they were more focused on their tobacco plantations. The same was true for the Catholic founders of Maryland. As their population grew, Protestants began to outnumber Catholics, though the Catholics continued to hold the power and influence. Just as in the other southern colonies, religion eventually took a back seat to tobacco in Maryland. Politics in the colonies were as different as their religious preferences. Seventeenth century New England was governed by Puritans. They governed the colonies with a General Court. The southern colonies, like Virginia, were ruled by the House of Burgesses. A large majority of the southern colonies followed the Virginia model of government. Northern colonial society was based on the Puritan religion. Community leaders attempted to form a completely pious society and to eliminate sin from within its boundaries. Those that did not conform were cast out or met with ridicule and violence, as is evident by the Salem witch trials. Southern colonists were mostly farmers with small plots of land that were maintained by family members and possibly a couple servants. Eventually, the mortality rate in the colonies began to decrease and most indentured servants survived long enough to be free. This caused a class system to develop that polarized the social structure of the south. Although they shared many similarities, and the prompt suggests that the Northern and Southern colonies were more similar than different, it seems clear that there were also many difference between the two. They had religious differences, they had political differences, they had economic and social differences. These differences prove the validity of the prompt’s statement to be wrong.