How to Overcome Insecurity in the Workplace

How to Overcome Insecurity in the Workplace Workplace insecurity is a thing- even for people who wouldn’t normally consider themselves â€Å"insecure.† Particularly for women, feeling less than on the job is a constant struggle- and usually, the feelings aren’t even warranted. Help yourself diagnose the most common workplace insecurities and figure out how you can overcome them. You’ll be much more productive and fulfilled if you do.1. Lack of TalentOkay, so right away you can probably write this off. You got hired for this job, didn’t you? So you can’t possibly be as underqualified or untalented as you think. Take a step back and stop comparing yourself and your work to that of your peers. Remember that your skill set and experience is unique and might bring something slightly different to the table than those of your coworkers- even if those might seem more impressive from where you’re sitting. And if you’re still feeling like you could know and do more†¦ learn a n ew skill or get a new qualification in an online course.2. Lack of AdvancementYou’re watching people get promoted all around you and you’re worried that you’re not advancing as fast as others. Don’t let the envy or resentment pull you down. Focus on your work. Sit down with your boss and have a chat about what you need to do to grow and start moving forward. Get clear on your expectations and then figure out how to exceed them!3. Lack of MoneyYou’re not supposed to talk about money in the office environment, but chances are you’ve been observant enough to know (ballpark) what the people around you are making. If this makes you feel inadequate, remember to make the focus on you: what could you be doing to qualify for a raise? Talk to your boss. Look around for other jobs, especially if you feel you’re not being rewarded for your skillset and work level.4. Lack of PopularityIf you feel your social skills aren’t quite up to par, s tart stepping up. Ask how you can be of help to coworkers. Go the extra mile. They will remember when the tables are turned and you need help. Plus, you can use their gratitude to build rapport and a better, warmer relationship.If you’re really stuck, there are lots of resources out there for improving your public speaking and social skills. Make use of them! And if you feel invisible on the job, start looking for ways to make yourself stand out a bit more. Take on high profile projects, come early, stay late, etc. A few calculated risks can put you in a much better position to be noticed and then valued.Remember that everyone makes mistakes and everyone is afraid of getting fired- at least at some point. Show up on time, do your job well, present yourself personably and professionally, and you should be in good stead. When in doubt, it’s always a good idea to solicit constructive feedback. Ask questions. Find allies. Trust your gut instincts.But more than anything: do your job and do it well. Go above and beyond. Rise above. There’s no better way to combat your insecurity than to achieve beyond even your own expectations. Just remember to take a proper moment to celebrate those achievements every time they occur.

Criminal law Essay Example | Topics and Well Written Essays - 2000 words

Criminal law - Essay Example The most important thing that needs to be point out in respect of criminal law is that it is based upon actus reus, mens rea and the absence of defences. The element of a reasonable person is required in respect of mens rea and therefore the other element that are required for the offences would not be discussed. Mens rea has been defined as a state of mind, intention or recklessness which is required for a certain crime. There is an ongoing debate and argument in respect of subjectivists and objectivists who tend to argue in different directions. The subjectivists argue that to the minimal level of serious crimes, the mental element that should be required should be awareness of the person’s actions and perceiving the surrounding circumstances as well as the results that would flow from the actus reus. On the contrary, the objectivists the argument rests on the fact that it should be sufficient mens rea if it is proved that a reasonable person would have foreseen the circumstances as well as the results that would flow from the actus reus and would therefore not be dependent on the awareness of the defendant. There have been competing claims which have been put forward by both. In respect of the subjectivists the main argument vests on the fact that there was self governance on the part of the individual who committed the crime and went contrary to the law. The objectivists on the other hand pointed out to the fact that there should be criminal punishment if it is found that his inadvertence was in respect of a substantial and obvious risk of the proscribed harm, which should have been perceived by the defendant. Even though there have been academic opinion that ‘the torch of orthodox subjectivism carried by Glanville Williams and Smith and Hogan and then by the Law Commission should be douse’ (Ashworth, POCL, p.253) it is still what is being followed in respect of serious offences and has been reiterated in the recent case of G1 where Lord Bingham said ‘it is a salutary principle that conviction of serious crime should depend on proof not simply that the defendant caused (by act or omission) an injurious result to another but that his st ate of mind when so acting was culpable. This after all, is the meaning of the familiar rule actus non facit reum nisi mens sit rea. The most obviously culpable state of mind is no doubt an intention to cause the injurious result, but knowing disregard of an appreciated and unacceptable risk of causing an injurious result or a deliberate closing of the mind to such risk would be readily accepted as culpable also. It is clearly blameworthy to do something involving a risk of injury to another...’ Despite of such an obvious favour being given to the subjectivist approach, there have been objectivist elements in respect of serious offences that have been laid down expressly by House of Parliament in particular sexual offences and certain offences related to money laundering. The element of intention are central to mens rea, however, in respect of the current situation the element of recklessness would be discussed. The situation in respect of criminal law has been that for crime s it was either intention or recklessness that would lead to construal of imposition of liability. In respect of recklessness there are two main elements that need to be construed, the first one being that of subjective recklessness and the other is that of objective recklessness. For establishing subjective recklessness the test that was laid down in Cunningham2 needs to fulfilled, which basically requires that an unjustifiable risk was taken as well as proving the fact that the defendant

Understanding Challenges In The Third Sector Essay - 1

Understanding Challenges In The Third Sector - Essay Example Hence, the third sector became a really interesting model for me to study and I learned how the entity succeed to survive when I worked as a volunteer with them. The organization I opted for this portfolio is a community organization St. Elizabeth’s, which manages children, youngsters, and adults within its school, college, and residential care home respectively. While becoming part of St. Elizabeth’s I realized that they work to support the needy with their problems. They were supported by charitable works and people like me volunteered there, hence, making it part of the third sector. Moreover, it also positively support the economy by providing employments and working opportunities to individuals like me who want to return something to their society. St Elizabeth’s is a traditional organization which was established in 1903 and has a history of more than 100 years. I learned that organization first charitable work was done by the Congregation of the Daughters of the Cross of Liege (the Congregation). During my volunteer work I also realized that they were a Roman Catholic religion based congregation, which was founded in 1833. However, over the period of time St Elizabeth’s has evolved and managed superior developments which cater to vulnerable people with disorders. Every institute operates to fulfill the goals, similarly, I comprehended that St Elizabeth’s also has its organizational goals and they focused towards achieving them. While working there as a volunteer; I noticed that the goal is to share care and compassion with every individual which require it the most. Hence, St Elizabeth’s was treating as much people as possible to overcome their learning disabilities. It plays a strong role as a NGO and has the support from the public sector. I think that there should be more organizations like St. Elizabeth’s working for the betterment of people with learning

Constrained Women Essay Example | Topics and Well Written Essays - 1000 words

Constrained Women - Essay Example Short stories can often focus a reader’s attention on various elements of our lives that frequently pass by under our radar because they are now so deeply ingrained in our society’s norms. Such concepts in our life would include the inherent male-dominant beliefs of old in which the woman’s space is considered to be only within the home. On a social level, the woman’s place remains below men even in modern times and is usually placed only slightly above children in matters of importance. While these are ideologies that have, in the past 100 years, been increasingly challenged, particularly by women who have continued to demonstrate their ability to keep up with men, it remains the case that women continue to struggle, not simply on the social level, but on the personal level as well. This personal battle in the example of female subservience requires women to begin questioning the values learned in their youth to consider something both thrilling and frigh tening – the idea that they may be capable of making their own decisions in life without the leadership or approval of men. For many women, the concept alone would be the most they would achieve, especially in earlier time periods. The female characters found in William Faulkner’s short story â€Å"A Rose for Emily† and Shirley Jackson’s short story â€Å"The Lottery† live within this oppressive ideology of the subservient woman both socially and internally, but also exhibit a yearning to escape the ties that bind them.

Respiratory Syncytial Virus (RSV) Vaccine Formulation

Respiratory Syncytial Virus (RSV) Vaccine Formulation Abstract Respiratory syncytial virus (RSV) is the leading cause of upper and lower respiratory tract infections in both children and elderly people. In this study we evaluated the short- and long-term protective efficacy of the single intranasal immunization of RSV vaccine formulation consisting of the fusion (F) protein formulated with a polyIC, an innate defense regulator peptide and a polyphosphazene (ΆF/TriAdj). This vaccine induced significant systemic and local immunity, including RSV F-specific IgG1 and IgG2a, IgA and virus neutralizing antibodies, and IFN-ÃŽÂ ³-secreting T cells in mice. Furthermore, this vaccination promoted the induction of RSV F85-93-specific CD8+ effector T cells with no virus replication in the lungs. To evaluate the duration of immunity induced by single intranasal vaccination, mice were challenged with RSV after five months of immunization. Significantly higher levels of IgG1, IgG2a and virus neutralizing antibodies were detected in ΆF/Tr iAdj vaccinated animals. Moreover, this vaccine formulation induced high local IgA secreting memory B cell development and B cell IgA production and conferred complete protection against subsequent RSV challenge without priming for enhanced disease. In conclusion, a single nasal vaccination of RSV ΆF protein formulated with TriAdj enhanced robust, long- term protective immune responses against RSV infection. Introduction Respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract disease in infants, older and immunocompromised individuals worldwide {Falsey, 2005 #586}. Many RSV vaccine candidates, including live attenuated strains, vectored and viral protein subunit vaccines are underway. Currently, there is no licensed RSV vaccine or specific treatments other than prophylaxis with passive antibody therapy (Palivizumab) {Group, 1998 #599}. The first RSV vaccine clinical trial with formalin-inactivated RSV (FI-RSV) failed to induce neutralizing antibodies and caused enhanced respiratory disease after natural RSV infection {Kim, 1969 #589}. RSV is an enveloped virus, containing three transmembrane surface glycoproteins: the major attachment protein G, fusion protein F and small hydrophobic SH protein. The F protein is highly conserved and mediates fusion of the virus into host cells and subsequent formation of syncytia, making it a major target for subunit vaccine and antiviral drug development {Collins, 2008 #597}. However, mucosal vaccination with RSV F protein alone does not induce protective immunity {Vaux-Peretz, 1992 #648;Garlapati, 2012 #650} therefore, this protein needs to be combined with an adjuvant to induce strong, long-lasting protective immune responses. Previously, we developed a novel combination adjuvant platform (TriAdj) comprising of three components, namely a polyI:C, innate defense regulator peptide (IDRs) and polyphosphazene (PCEP). Synthetic double-stranded RNA, polyIC, known to recognize TLR3 and retinoic acid-inducible gene 1(RIG-I) leading to induction of proinflammatory cytokines, which in turn activates various immune cells {Sivori, 2004 #581`, Longhi`, 2009 `#573;Longhi, 2009 #573}. To stabilize and protect the polyIC from degradation, two other immunmodulators an innate defense regulator (IDR) peptide and a polyphosphazene were added. Innate defense regulator (IDR) peptides are derivatives of natural host defense peptides with microbicidal, chemotactic properties {Yeung, 2011 #608}. Poly[di(sodium carboxylatoethylphenoxy)]-phosphazene (PCEP) is a synthetic biodegradable water-soluble polymer with immunostimulatory properties. It forms non-covalent complexes with antigens and/or other adjuvants to increase their stabil ity and allow multimeric presentation {Andrianov, 2009 #618;Kovacs-Nolan, 2009 #621}. Previously, we developed a novel vaccine candidate against RSV (ΆF/TriAdj) consisting of a truncated version of the RSV fusion protein (ΆF) formulated with TriAdj. We have shown that double vaccination regime such as prime and boost intranasal immunization with this vaccine candidate establishes long-lasting humoral and cell-mediated protective immune responses without any evidence of immunopathology {Garg, 2014 #60;Garg, 2015 #63}{Garg, 2014 #60}{Garg, 2014 #60}{Garg, 2014 #60}{Garg, 2014 #60}{Garg, 2014 #60}. In the present study we demonstrated that after single IN delivery with ΆF/TriAdj, vaccine elicits mucosal and systemic immune responses and offers complete protection from RSV challenge in mice. Furthermore, ΆF/TriAdj induced effective local CD8+ T cells, which is one of the hallmarks for successful vaccination against many viral infections. To evaluate the duration of immunity induced by this vaccine candidate after single intranasal vaccination, we carried out long-term trials. Five month after the vaccination, RSV ΆF formulated with TriAdj induced robust mucosal and systemic immune responses and complete protection from RSV challenge in mice. Furthermore, ΆF/TriAdj elicited long-lived local IgA secreting memory B cell development and B cell IgA production, as well as memory T cells. It is well known that an RSV vaccine needs to induce a balanced Th1/Th2 adaptive immune response, which stimulates the generation of high-affinity neutralizing antibodies with effector cytotoxic CD8+ T cells {Graham, 2011 #642}. Results Single mucosal vaccination with ΆF/TriAdj induces strong systemic immune responses: Antibodies play a major role in protection against many viruses. To characterize the quality of the humoral immune responses induced by single vaccination of ΆF/TriAdj, IgG1 and IgG2a titers were measured in the serum. As shown in Fig. 1A B, the mice immunized with ΆF/TriAdj developed significantly higher IgG1 and IgG2a levels, than placebo groups both before and after challenge with RSV. These results indicate that formulation of ΆF protein with TriAdj promotes a balanced humoral immune response. Neutralizing antibody plays a protective role against viral infections. In order to evaluate the biological function of the ΆF-specific serum antibodies, Virus neutralization (VN) titers were determined. Mice immunized with ΆF/TriAdj developed significantly higher neutralizing antibody titers than the ones immunized with PBS both before and after challenge with RSV (Fig 1C). To investigate the phenotype of the cellular immune response, the ΆF-induced secretion of IFN-ÃŽÂ ³ and IL-5 by splenocytes was measured four days after challenge. Significantly higher numbers of IFN-ÃŽÂ ³ secreting cells were generated from mice vaccinated with ΆF/TriAdj formulation, while the number of IL-5 secreting cells was low in vaccinated mice (Fig 1D). This suggests the addition of RSVΆF protein to the TriAdj resulted in a stronger, Th1-biased immune response, which is in also agreement with the enhanced IgG2a production. Intranasal vaccination with single dose of ΆF protein formulated with TriAdj induces robust protective mucosal immune: Mucosal immune response play a crucial role in first line defense of host by blocking the spreading of RSV infection in the lungs. Thus, an effective RSV vaccine should induce local IgA antibodies in the respiratory tract. To evaluate the mucosal immune responses induced by ΆF/TriAdj, we measured ΆF-specific IgA antibody levels in the lungs. Significantly higher IgA production was observed in mice immunized with ΆF/TriAdj in comparison with mice immunized with PBS (Fig 2A). To determine whether single IN vaccination with the ΆF/Triadj formulations would affect viral clearance, all mice except those in one of the two Placebo groups were challenged IN with RSV. No infectious virus was recovered in lungs of mice immunized with ΆF/TriAdj, showing full protection from infection (Fig 2B). In order to investigate whether protection is correlated with higher cross-presentation and cell-mediated immune responses, we evaluated RSV F-specific CD8+ T cells infiltration with RSV F85-93 pentamers in the lungs after RSV challenge. The ΆF/TriAdj induced a significantly higher number of F85-93 specific CD8+ T cells influx in the lungs (Fig 6A), which suggests that vaccination with ΆF/TriAdj promotes a cytolytic CD8+ T cell response to RSV infection. Virus-specific effector CD8+ T-cell response has been shown to play a critical role in RSV clearance {Graham, 2011 #642}. Therefore, we analyzed the effecter function of the CD8+ T cells according to their expression of the IFN-ÃŽÂ ³ by à ¯Ã‚ ¬Ã¢â‚¬Å¡ow cytometry. After RSV challenge, the ΆF/TriAdj-immunized group showed higher frequencies of IFN-ÃŽÂ ³-expressing CD8+ T cells than PBS-immunized, RSV-challenged mice in the lungs (Fig 6). This suggests that ΆF/TriAdj promotes a stronger ef fector CD8+ T cell response to RSV. Single nasal vaccination with ΆF/TriAdj induces Long-term systemic immune responses: To assess the long term memory effect induced by single intranasal vaccination with ΆF/TriAdj, serum IgG and VN antibody levels were measured periodically over five months. The mice immunized with ΆF/TriAdj developed strong IgG and VN titers compare to placebo control group for approximately 25 weeks (Fig 3A B). We next examined whether IN vaccination promoted the induction of long term ΆF-specific serum IgG1 and IgG2a. The ΆF/TriAdj- vaccinated mice developed significantly higher ΆF-specific serum IgG1 and IgG2a titers than placebo groups both before and after challenge with RSV (Fig 4A B). VN titers were also determined to evaluate the functional quality of the ΆF-specific serum antibodies after RSV challenge. Mice immunized with ΆF/TriAdj developed significantly higher neutralizing antibody titers compared to PBS gro up (Fig 4C). To further define the phonotype of the immune response, the ΆF-induced secretion of IFN-ÃŽÂ ³ and IL-5 by splenocytes was measured four days after challenge with RSV. The ΆF/TriAdj formulation generated a significantly higher frequency of IFN-ÃŽÂ ³ secreting cells compared to the placebo. The numbers of IL-5 secreting cells were low in all vaccinated mice (Fig 4D), which suggests Th1 bias immune responses. Mucosal vaccination with Single dose of ΆF/TriAdj induces Long-term protective mucosal immune response: Local antibody play a major role in protection from respiratory viruses such as RSV in the lung. To access the long term local immune responses with ΆF/TriAdj vaccination, induction of ΆF-specific IgA in the lungs was examined by ELISA. Mice immunized with ΆF/TriAdj developed significantly higher IgA levels when compared to placebo and placebo virus challenged groups (Fig 5A). Furthermore, we checked the development of IgA-secreting memory B cells in lungs after RSV challenge. ΆF/TriAdj formulations generated a significantly higher frequency of RSV ΆF-specific IgA-secreting memory B cells in lungs in comparison to PBS (Fig 5B). In summary, single dose of ΆF/TriAdj via IN vaccination stimulated strong IgA-secreting memory B cell development and B cell IgA production. To determine viral clearance after single vaccination with ΆF/TriAdj, all mice except those in one of the two placebo groups were challenged IN with RSV on day 150. The mice were sacrificed after four days, and virus titers in the lungs were determined. No infectious virus particle was recovered from mice vaccinated with ΆF/TriAdj, showing that these mice still had sufficient immunity to be completely protected from challenge virus replication in the lungs (Fig 5C). Discussion RSV is one of the major global burdens of causing a broad spectrum of respiratory illnesses in children and older population worldwide. There is still no licensed RSV vaccine, in part due to the disastrous outcomes observed following vaccination of naÃÆ' ¯ve children with FI-RSV {Kim, 1969 #589}. Natural infection with RSV fails to protect against subsequent infection because it stimulate modest immunogenicity and short-lived immunological memory against virus. Similar to natural RSV infection, many promising RSV vaccine have failed to generate long-lasting, protective immune responses {Power, 2008 #769;Hall, 2001 #789}. For wide clinical application, RSV vaccine will need to be safe, easy to administer and must stimulate strong long-term protective immunity than natural RSV infection {Pulendran, 2011 #796}. Protein subunit vaccines have a high safety profile, but are generally poorly immunogenic and induce short-lived humoral and cellular immunity {Graham, 2011 #642}. So adjuvant s are added to subunit vaccines to stimulate faster, stronger, and long-lasting immune responses to vaccines. At the clinical level, multi dose vaccination is expensive and has potential safety problems. Whereas, single vaccination formats are cost effective, safe (no risk for vaccine contamination) and more convenient to health worker. In the present study, we demonstrated the short as well as long term protective efficacy including stable systemic IgG, local IgA production and neutralizing antibody titers of a RSV ΆF protein formulated with TriAdj in mice after single IN immunization. Furthermore, ΆF/TriAdj promoted an effector CD8+ T cells with compete protection against RSV infection without inducing vaccine-enhanced pathology in the lungs. A vaccine requiring only a single administration to be effective would be highly practical, as it would improve patient compliance Therefore, single intranasal immunization with RSV ΆF protein formulated with Tri Adj is a promising approach to induce a long-lasting protective RSV-specific immune response. Mucosal immunization via IN route is invasive and suitable for generation of both local and systemic immunity. RSV infect through respiratory tract, therefore mucosal antibodies specifically IgA play an important role in prevention and clearance of virus. Previously it has been shown that protection against RSV infection is more correlated to the levels of mucosal IgA rather than to systemic antibody in human {Watt, 1990 #687}. In mice, single RSV infection induces short-lived neutralizing antibodies with absence of antibody-secreting memory B cells {Schmidt, 2012 #869}. Interestingly, we have recently demonstrated that the RSV infected mice were showed significantly lower level of local B cell IgA production and IgA secreting memory B cell development that indicates an impaired local antibody response that allows RSV re-infection and explain the short-term protective immunity with natural infection with RSV{Garg, 2016 #1340}. In contrast, intranasal vaccination with single dose of R SV ΆF protein formulated with TriAdj promotes the induction of long-lasting local IgA secreting memory B cell development and B cell IgA production which is a major criterion for an effective RSV vaccine. Previously, we and others have reported that strong, effective and protective immune responses were induced only after a booster immunization with RSV F protein with adjuvant {Garg, 2014 #891}{Blanco, 2014 #1423}{Lambert, 2015 #1442}. A single mucosal vaccination of adenovirus-based RSV vaccine had also been proved effective in short term {Kim, 2010 #1443}. But this is the first report showing the long term protective efficacy of single mucosal vaccination of an adjuvanted RSV F protein subunit vaccine in mice. Clinical trial with FI-RSV failed to induce protection against RSV due to generation of poor neutralizing antibodies and TLR activation that led to vaccine-associated enhanced disease {Delgado, 2009 #513}. The inclusion of polyI:C as TLR ligand in vaccine formulation, is expected to overcome the limitations of inactivated RSV vaccines. The adjuvant effect such as long-lasting T cell immunity of polyI:C is likely caused by the direct interaction with PRRs such as TLR3, MDA5 and RIG-I, leading to production of pro-inflammatory cytokines, and chemokines. Interestingly, it has been shown that alveolar macrophages and lung DCs induced strong IgA and IgG antibodies by addition of TLR3 ligands, which were probably linked to secretion of BAFF/APRIL cytokines and activated B cells {Xu, 2008 #686}. However, we and others have been shown that IN administration of higher doses of polyIC caused marked production of inflammatory cytokines/chemokines accompanied by impaired lung function {Stowell, 2009 #682;Boukhvalova, 2010 #685;Aeffner, 2011 #750} suggesting excessive stimulation of local immune responses can result in detrimental effects. In contrast, ΆF/TriAdj has an advantage over polyIC alone as it has shown full protection form RSV infection without any exacerbation of chronic pulmonary disease. This demonstrates that a combination of polyIC, IDR and PCEP mediates optimal enhancement of the RSV-specific immune response without vaccine enhanced disease. We believe that IDRs is one of the major anti-inflammatory components in our vaccine formulation as it has been previously shown to enhance cell-mediated immune responses and to modulate excessive consequences of TLR signaling {Bowdish, 2005 #766;Hancock, 2006 #767}, whereas polyphosphazenes enhance antigen-specific humoral immunity through the formation of non-covalent complexes with protein {Andrianov, 2005 #667}, suggesting major roles from both of these compounds in this vaccine formulation. An important factor for successful vaccine against RSV requires the generation of effective CD8+ T cell in the lungs. Previously, it has been shown that mild to severe infection with RSV failed to induce virus-specific IFN-ÃŽÂ ³ recall responses in the infants {Lee, 2007 #806}. To define the stimulation of local effective CD8+ T cell responses induced by ΆF/TriAdj, RSV F specific pentamer staining demonstrated marked induction of IFN-ÃŽÂ ³ secreting RSV-specific CD8+ T cells in lungs. Similarly, exogenous IFN-ÃŽÂ ³ expression protects against RSV infection in the lungs of BALB/c mice {Kumar, 1999 #805}. This evidence suggests that complete protection against RSV was associated with the presence of IFN-ÃŽÂ ³+ RSV-specific CD8+ T cells. In summary, our data show that RSV ΆF protein formulated with TriAdj vaccine represents a safe, effective and promising RSV vaccine candidate. The major advantages associated with this mucosal RSV vaccine is to induction of long-term protective immunity with a single mucosal vaccination by stimulating long-lived RSV-specific neutralizing antibodies, memory B and CD8+ T cells, therefore warranting additional evaluation as a vaccine against RSV in clinical trials. MATERIALS AND METHODS Virus and vaccine formulation: The RSV (A2 strain) was propagated in Hep-2 cells (American Type Culture Collection, VR-1540). The RSV ΆF protein with his-tag was produced and purified as described previously {Garlapati, 2012 #650}. Briefly, HEK-293T cells were transfected with an episomal vector expressing the ΆF protein using Turbofect (Fermentas, R0534). The ΆF protein with his-tag was purified using TALON Superflow resin (Clontech,) according to the manufacturers instructions. The ΆF protein was formulated with 10  µg poly (I:C) (Invivogen, tlrl-picw), 20  µg IDR1002 (Genscript, 818360) and 10  µg PCEP (Idaho National Laboratory) in PBS (ΆF/TriAdj) as described previously. Animals, immunizations and challenge: Six to eight week old female BALB/c mice (Charles River Laboratories, Wilmington, MA, USA) were immunized once IN with ΆF/TriAdj. Two additional groups of mice received PBS IN (Placebo control). Three week (short-term trial) and twenty weeks (long term trial) post-vaccination, all except one of the Placebo groups were challenged IN with RSV strain A2 (5 x 105 PFU/50 µl). Mice were euthanized four days after RSV challenge for detection of immune response and virus in the lungs. Blood samples were drawn by cardiac puncture under anesthesia prior to challenge and at regular intervals afterwards. All procedures were approved by the University Animal Ethics Committee in accordance with the standards stipulated by the Canadian Council on Animal Care. Lung fragment cultures and ELISA: Lavaged lungs of euthanized mice were cut into small pieces and lung fragment cultures were prepared as described previously {Mapletoft, 2008 #591}. ELISA was performed as described previously {Garg, 2014 #942}. RSV ΆF-specific IgG1, IgG2a and IgA were detected using biotin-labeled goat anti-mouse IgG1, IgG2a (Southern Biotech, 1070-08, 1080-08) or IgA (Gibco, M3115) followed by streptavidin-alkaline phosphatase (AP) (Jackson ImmunoResearch Laboratories Inc.,016-050-084), and developed with p-nitrophenyl phosphate (Sigma-Aldrich, N3254) substrate. Enzyme-linked immunospot (ELISPOT) assays: ELISPOT assays were performed on splenocytes and LNs cells as previously described {Garlapati, 2012 #650}. Briefly, cells were stimulated with ΆF protein (2  µg/ml) and spots were developed using biotinylated IFN-ÃŽÂ ³- or IL-5- (BD Biosciences, 554410, 554397) or goat anti-mouse IgA specific antibody, AP-conjugated streptavidin and BCIP/NBT (Sigma-Aldrich, B5655) as the substrate. Analysis of CD8+ T cells from lungs by flow cytometry: Lung single-cell suspensions were generated as previously described {Garg, 2014 #942}. To analysis of RSV F85-93-specific CD8+ T cells, lung mononuclear cells were surface stained with H-2Kd-F85-93 MHC Class I pentamer (ProImmune, F149) together with labeled antibodies specific for CD8+ T cells (BD Pharmingen, 553031). For intracellular detection of IFN-ÃŽÂ ³ in CD8+ T cells, lung mononuclear cells were stimulated with 1 ÃŽÂ ¼M of F85-93 peptide (KYKNAVTEL) and cell surface staining was performed, followed by intracellular cytokine staining using Cytofix/Cytoperm (BD Pharmingen, 554715) and APC- conjugated anti-IFN-ÃŽÂ ³ antibody (BD Biosciences, 554413). After staining, cells were acquired by flow cytometry (BD Biosciences), and data were analyzed using Kaluza Software (Version 1.2). Cells were gated for live cells, singlets and lymphocytes and then analyzed for indicated markers. Virus titration and virus neutralization assay: Virus titrations were performed with individual lungs at day 4 post- challenge as described previously {Garg, 2014 #942}. Results are expressed as PFU/g of lung tissue. RSV-specific neutralization titers were determined by plaque reduction assays. Serum samples were mixed with 500 PFU/well of RSV strain A2 for 1 h at 37 °C. The sample-virus mixtures were transferred to HEp-2 cell monolayers and incubated for 4 days at 37  °C, and the cells were fixed and stained with 0.5% crystal violet. Statistical analysis: All data were analyzed using GraphPad PRISM version 6 for Windows (GraphPad Software, Inc., La Jolla, CA, USA). Differences among all groups were examined using Student t-tests, one-way ANOVA, followed by a Newman-Keuls post test. Differences were considered significant if P Acknowledgements The authors would like to thank Elisa C. Martinez, Wayne Connor, and Michael Theaker for technical assistance, as well as Sherry Tetland and Jan Erickson for the handling and care of the animals. This work was supported by the Krembil Foundation, and the Canadian Institutes of Health Research (CIHR). Published as VIDO manuscript number . Figure legends Fig. 1. RSV ΆF-specific systemic humoral immune responses in mice. (A) Serum IgG1 and IgG2a titers before challenge (B/C), (B) Serum IgG1 and IgG2a titers after challenge (A/C) with RSV, (C) Serum VN antibody titers determined before (B/C) and after (A/C) RSV challenge, and (D) Numbers of IFN-ÃŽÂ ³ and IL-5 secreting splenocytes determined in response to in vitro restimulation with ΆF protein. BALB/c mice were immunized once IN with ΆF formulated with TriAdj and challenged three weeks later with RSV. Control groups were immunized with PBS and challenged with RSV (Placebo) or mock-challenged (Placebo/mock). ELISA titers are expressed as the reciprocal of the highest dilution resulting in a value of two standard deviations above the negative control serum. Virus neutralization titers are expressed as the highest dilution of serum that resulted in Fig. 2. Mucosal immune responses to RSV ΆF protein in mice after challenge with RSV. The IgA titer (A), Virus titer (B), Percentages of ΆF-specific CD8+ T cells (C), and ΆF-specific IFN-ÃŽÂ ³ secreting CD8+ T cells (D) were measured in the lung after RSV challenge. Mice were immunized and challenged as described in the legend for Fig. 1. ELISA titers are expressed as the reciprocal of the highest dilution resulting in a value of two standard deviations above the negative control serum. Virus replication in the lungs is expressed as pfu per gram of lung tissue. Bars represent median values with interquartile ranges. *P Fig 3. Long-term systemic immune responses to RSV ΆF protein. IgG (A) and VN (B) titers were measured at different times after vaccination. BALB/c mice were immunized once IN with ΆF formulated with TriAdj and challenged with RSV on day 150. Control groups were immunized with PBS and challenged with RSV (Placebo) or mock-challenged (Placebo/mock). ELISA titers are expressed as the reciprocal of the highest dilution resulting in a value of two standard deviations above the negative control serum. Virus neutralization titers are expressed as the highest dilution of serum that resulted in Fig 4. Systemic immune responses to RSV ΆF protein. (A) Serum IgG1 and IgG2a titers before challenge (B/C), (B) Serum IgG1 and IgG2a titers after challenge (A/C) with RSV, (C) Serum VN antibody titers determined after (A/C) RSV challenge, and (D) Numbers of IFN-ÃŽÂ ³ and IL-5 secreting splenocytes determined in response to in vitro restimulation with ΆF protein. Mice were immunized and challenged with RSV as described in the legend for Fig. 3. ELISA titers are expressed as the reciprocal of the highest dilution resulting in a value of two standard deviations above the negative control serum. Virus neutralization titers are expressed as the highest dilution of serum that resulted in Fig 5. Long term mucosal immune responses to RSV ΆF protein. IgA titers (A), numbers of IgA secreting LNs cells (B), Virus titers (C) were determined after (A/C) RSV challenge. Mice were immunized and challenged with RSV as described in the legend for Fig. 3. ELISA titers are expressed as the reciprocal of the highest dilution resulting in a value of two standard deviations above the negative control serum. IgA secreting cell numbers are expressed as the difference in the number of spots between ΆF-stimulated wells and medium-control wells. Virus replication in the lungs is expressed as pfu per gram of lung tissue. Bars represent median values with interquartile ranges. **P

Emily dickinson Essay examples -- essays research papers

Emily Dickinson’s poetry powerfully indicates values of society of the time. It does this through its conciseness, its simplicity and its control. Indications of society’s values are seen in many of Dickinson's poems, but they are especially noticeable in ‘It was not Death’, and ‘Because I could not stop for Death’. In Dickinson’s poem ‘It was not Death’, she demonstrates how restricting and stereotyping society can be on an individual, and how society values the conformity of the whole community, even though they may not want to. In Dickinson’s poem ‘Because I could not stop for Death’, she is questioning society’s values on religion and everlasting life. Emily Dickinson’s poems analyze her perception of the world and society, which is different to that of the commonly accepted, objective perception. The reader sees this perception in her poem ‘It was not Death’, where Emily appears to perceive a world full of confusion and chaos. She also observes that society tries to place people into stereotypes, and feels that she herself is restricted to one. The Figures I have seen Set orderly, for Burial, Reminded me, of mine – Dickinson shows in these lines that her own life reflects that of a dead persons – it appears to be a living thing, but lacks something that makes it alive. It seems that life is a convential pattern, and she is conformed in society just like the people in the coffins. She resents the way that in her society people were heavily placed into stereotypes. As if my life were shaven, And fitted to a frame These lines express Dickinson’s thoughts about the restrictions of her life in her society. The fact that her life was ‘shaven’ seems to give the image of being cut down to size with a razor to fit her frame, and this is a very sharp image. It also seems to hold connotations to the times of torture and the methods they used, and she may be suggesting that the rest of society make her life torture. It is as if her whole life has been shaped and trapped, which is not by its own nature, and from which it can not escape. Emily Dickinson also gives the impression of confusion and chaos through the verse techniques employed in her poem ‘It was not Death’. There are a mixture of images which give the impression of confusion and chaos. In the poem there is action (‘I stood up), sound (‘Bells’), frost, heat (‘Siroccos’, ‘Fire’), shipwreck (‘Without a Ch... ...else, it just simply stops, whilst you are still aware of it. Since then – ’tis Centuries – and yet Feels shorter than the Day I first surmised the Horses’ Heads Were toward Eternity – The lack of punctuation in the last stanza is to show breathlessness and panic as the woman realises that she has come to a state of eternity, and also emphasises that this is a long time – forever. This is how Emily Dickinson shows her theory of eternity after death, which contradicts the values of the rest of society. Values of society of the time can be seen through Emily Dickinson’s poetry. She powerfully shows society’s values of conformity in her poem ‘It was not Death’, as well as how she and many others were expected to fit into stereotypes, even if it meant sacrificing their natural state. This is shown through the form of the poem and the poetic devices that are used. ‘Because I could not stop for Death’ powerfully shows some of society’s values by contradicting them. Emily Dickinson displays her own beliefs on what comes after death, and they are certainly not to do with an afterlife in Heaven, or other religious beliefs. These are examples of how Emily Dickinson’s poems are powerful in

English Literature Essay

Literature (from Latin litterae (plural); letter) is the art of written work, and is not confined to published sources (although, under some circumstances, unpublished sources can also be exempt). The word literature literally means â€Å"things made from letters† and the pars pro toto term â€Å"letters† is sometimes used to signify â€Å"literature,† as in the figures of speech â€Å"arts and letters† and â€Å"man of letters.† The four major classifications of literature are poetry, prose, fiction, and non-fiction. What in Philippine Literature? Philippine literature is the literature associated with the Philippines and includes the legends of prehistory, and the colonial legacy of the Philippines. Most of the notable literature of the Philippines was written during the Spanish period and the first half of the 20th century in Spanish language. Philippine literature is written in Spanish, English, Tagalog, and/or other native Philippine languages. As a Filipino, What is the importance of Literature? Literature serves as an enormous information base. Research works by famous inventors and literary works by notable scientists often narrate stories of their groundbreaking discoveries and inferences. Ongoing developments in the fields of science and technology are documented so that the world can know about them. Several ancient scriptures relating stories of human evolution and narratives of human life in those times, have been of tremendous help to mankind. Thus, literature has always served as an authentic source of information from all around the world. What environmental education goals and features? Philippines is not an exemption in letting us feel the harsh realities of these environmental threats and challenges. With all the ever increasing demands created by the environmental problems accompanied by social, political and technological changes happening, the education sector is faced with challenges and is expected to initiate changes in the curriculum and give more attention to environmental education. The curriculum should be geared towards the provision of knowledge, skills and values that will help  the learners cope and adapt to these changes. The goal of environmental education is to develop an environmentally literate and responsible citizenry who will ensure the protection and the improvement of the environment, and bring about sustainability, social equity and economic efficiency in the use of the country’s natural resources. Because of the existing environmental problems we are into and the thrust for sustainability, the protection and improvement of the environment have become imperative educational goals. It is also envisioned that within the next decade, a great majority of Filipinos will be imbued with a sense of responsibility to care for, protect for, and enhance environmental quality that is conducive to their wellbeing and supportive of the nation’s economic development.