Slutsteget i immunförsvarets komplementaktivering innefattar en klyvning av komplementprotein C5, till C5a och C5b, vilket initierar bildandet av terminalt komplementkomplex (TCC) som i form av membran-attack-komplex (MAC) bildar cytotoxiska porer i bland annat gramnegativa bakterier. Bildandet av MAC kan blockeras av endogena regulatorer och TCC frisätts då som ett lösligt komplex, sC5b-9, i plasma. I examensarbetet studerades en variant av TCC, som i tidigare studier visats bildas oberoende av C3- och C5-konvertas när serum surgjorts till pH < 7,0 in vitro. Syftet med studien var att studera om detta atypiska TCC (aTCC) bildades hos grisar, som i en mekonium aspirationssyndrom (MAS)-modell, erhållit ett sänkt systemiskt pH in vivo. I syftet ingick också att etablera en ELISA-baserad metod för att analysera aTCC. I en sandwich ELISA användes monoklonal anti-C5a/C5a (desArg) (klon T13/9) som fångande antikropp och monoklonal anti-C9 (klon aE11) som detekterande antikropp för att analysera aTCC i plasmaprover från 18 MAS-grisar, samt i ett kontrollmaterial bestående av grisserum som surgjorts till pH 6,8 och 6,4 in vitro. Mängden aTCC i kontrollproverna ökade när pH sänktes men innehållet av aTCC i plasmaproverna minskade över MAS-studiens förlopp. När den relativa förändringen i aTCC relaterades till MAS-grisarnas slutliga pH kunde ett signifikant samband ses (p = 0,02) som visade att en större förändring i aTCC sammanföll med ett lägre slutligt pH. Nivåerna av aTCC var generellt sett högre i plasmaproverna jämfört med kontrollproverna vilket skulle kunnat bero på skillnader i plasma vs serum avseende aTCC eller att proverna kom från grisar med olika ålder och vikt. Avsaknad av grisspecifik standard och negativ kontroll samt lågt signal/brusförhållande bidrar till felkällor för analysen och denna kräver fortsatt optimering. / The late steps of complement activation involves a cleavage of complement protein C5, to C5a and C5b, which initiates the formation of terminal complement complex (TCC). The final complex is referred to as the membrane-attack-complex (MAC) which forms cytotoxic pores in, inter alia, gram-negative bacteria. The formation of MAC can be inhibited by endogenous regulators and the TCC is then released as a soluble complex, sC5b-9, in plasma. In the degree project, another type of TCC was studied, which in previous studies had shown to form independently of C3 and C5 convertases when serum was acidified to pH <7.0 in vitro. The purpose of the study was to investigate whether this atypical TCC (aTCC) was formed in piglets, which in a model of meconium aspiration syndrome (MAS), received a reduced systemic pH in vivo. The purpose was also to establish an ELISA for analyzing aTCC. Sandwich ELISA, with monoclonal anti-C5a / C5a (desArg) (clone T13/9) as a capture antibody and monoclonal anti-C9 (clone aE11) as a detection antibody, was used to analyse aTCC in plasma samples from 18 MAS piglets, and in control samples consisting of pig serum acidified to pH 6.8 and 6.4 in vitro. The amount of aTCC in the control samples increased when the pH was lowered, but the content of aTCC in the plasma samples decreased over the course of the MAS study. When the relative change in aTCC was related to the final pH of the MAS pigs, a significant relationship could be seen (p = 0.02) which showed that a major change in the aTCC coincided with a lower final pH. aTCC were generally higher in plasma samples compared with control samples, which could be due to differences in plasma vs serum for aTCC or that the samples came from pigs of different age and weight. Lack of pig-specific standard and negative control as well as low signal to noise ratio contribute to sources of error for the analysis and this requires continued optimization.

Marine bacteria play an important role in the marine nutrition cycles. About half of all sea-living bacteria can use light as an energy source, in addition to organic carbon compounds, which is important for survival as the seas becomes acidic and low in nutrition due to changing climate. The Flavobacteriaceae is a bacterial family that plays an important role in the degradation of complex organic compounds in natural environments. Dokdonia donghaensis MED134 belongs to the Flavobacteriaceae family and use both chemotrophy and phototrophy as a source of energy. For its phototrophic ability, the bacteria use proteorhodopsin, which is a membrane-bound proton pump. This allows the bacteria to survive and grow in nutrient-poor environments. The purpose of the study was to investigate with qPCR if there was a difference in gene expression for proteorhodopsin and isocitrate dehydrogenase in Dokdonia donghaensis MED134 depending on whether the bacteria had grown in constant light or darkness. Analysis with qPCR showed a significantly greater gene expression for proteorhodopsin in the bacteria grown in constant light for seven days, compared to those grown in constant light for three and four days and those grown in constant darkness. No significant difference could be demonstrated in gene expression for isocitrate dehydrogenase. This indicates that Dokdonia donghaensis MED134 in nutritional deficiency can use light as a source of energy for survival and further growth. By simulating climate change an adaptability could be seen through increased gene expression. Through this, further understanding of the role of bacteria in the marine ecosystem can be obtained and further research can be conducted as the marine climate issue becomes more relevant.

Herpes simplexvirus 1 och 2 orsakar godartade sjukdomar, men kan även orsaka mortalitet. Diagnostik av herpes simplexvirus 1 och 2 utförs numera framför allt med realtids- Polymerase chain reaction (PCR). I metoden amplifieras specifika deoxiribonukleinsyra(DNA)-sekvenser till miljontals kopior vilka sedan detekteras med fluorescein. I realtids-PCR sätts positiva och negativa kontroller. De positiva kontrollerna kan vara InHouse eller kommersiella. Tolkningen av resultatet inkluderar inspektion av kontrollerna. DNA utsätts för nedbrytningsprocesser av olika slag, och kan förvaras på olika sätt för att upprätthålla stabilitet. Syftet med studien var att jämföra laboratoriets InHouse-kontroller med två kommersiella kontroller, för att utvärdera vilken av dessa som var mest stabila över tid. Utvärderingen utfördes genom att analysera tre kontroller med realtids-PCR, efter att de hade förvarats i -20° C, i 5° C och i 20° C, och var spädda i TE-buffert eller i PCR-vatten. De kommersiella och InHouse-kontrollerna visade sig vara jämbördiga. Vidare studier som görs under längre tid, i större omfattning och där koncentrationerna är samma för varje kontroll, föreslås. / Herpes simplex viruses 1 and 2 which usually cause benign diseases but can even cause mortality. The diagnostics of herpes simplex virus 1 and 2 are performed with real-time Polymerase chain reaction (PCR). In the real-time PCR method, specific deoxyribonucleic acid (DNA) sequences are amplified into millions of copies which are then detected with fluorescein. Positive and negative controls are used in real-time PCR. The positive controls can be InHouse or commercial. The interpretation of the results includes inspection of the controls. DNA is subject to degradation processes of different kinds and can be stored in different ways to maintain stability. The purpose of the study was to compare the laboratory's InHouse controls with two commercial controls, to evaluate which of these were more stable over time. The evaluation was performed by analyzing the three controls with real-time PCR after they were stored in temperatures at -20° C, at 5° C and at 20° C, and were diluted in TE-buffer or in water. The commercial and InHouse controls proved to be equitable. Further studies carried out for a longer period of time, to a greater extent and where concentrations are the same for each control are suggested.

Forcerad exspiratorisk volym på en sekund (FEV1) och vitalkapacitet (VC) utgör grunden för spirometri som är ett diagnostiskt hjälpmedel vid lungsjukdomar. Spirometri utförs vanligtvis i sittande position, men kan utföras i stående position. Syftet med studien var att jämföra om det finns en signifikant skillnad för FEV1 och VC vid dynamisk spirometri mellan sittande och stående position hos studenter utan känd lungsjukdom. Datainsamlingen utfördes på Klinisk Fysiologi, Länssjukhuset Ryhov i Jönköping av legitimerad biomedicinsk analytiker. 13 frivilliga studenter i åldrarna 22-33 deltog i studien, fyra var män och nio var kvinnor. Genomsnittligt BMI var 21,9 kg/m2 . Manövrarna utfördes minst tre gånger i sittande och sedan stående position. Deltagare med längd över 175 cm fick stå på knä. Medianen för VC i sittande position var 4,5 liter respektive 4,4 liter i stående position. Medianen för FEV1 var 3,6 liter i samtliga kroppspositioner. Wilcoxon-rangsummetest påvisade ingen statistiskt signifikant skillnad för varken VC eller FEV1 mellan sittande och stående position. På grund av litet urval kan ingen generell slutsats dras av denna studie men kan utgöra underlag för fortsatta studier. Ytterligare studier med en större och mer spridd population krävs för att kunna dra generella slutsatser om kroppspositionens påverkan på FEV1 och VC.

The result of cryopreservation of semen is crucial for patients in need of fertility preservation.The cryopreservation method is not optimized since only 10 % the sperms are expected tosurvive the treatment. The sperms are exposed to many risk factors such as oxidative stress,osmotic chock and ice crystallization. To minimize the risks, use of cryoprotectants is needed.The use of cryoprotectants helps the cell to dehydrate as penetrating cryoprotectants cancreate space between ice crystals and cell membrane.Two studies were performed. In study one, two different freezing medias (SpermFreezesolution and Cryoprotect II effect on sperm motility after freezing in were compared). Studytwo investigate whether the motility were best preserved if semen froze with all the contentsof the ejaculate or if the sample should be concentrated, with removal of seminal plasma,epithelium cells and dead sperm cells by gradient centrifugation.The project was performed according to recommended instructions for each freezing media.In total, 55 samples were collected for the first study and 23 samples were collected for thesecond study. The sperm motility was measured both before freezing and after thawing.The Cryoprotect II medium preserved the cells better than the SpermFreeze medium(p=0,006). Using SpermFreeze, higher rate of motility was obtained when centrifugation wereperformed before freezing (p=0,033), while this was not observed when using Cryoprotect II(p=0,055). In conclusion Nidacon preserved sperm more effectively than Vitrolife freezemedium. Vitrolife’s freezing medium preserved the samples better if centrifugation wereperformed before freezing. Nidacons freezing medium gave the same result for the samplesno matter centrifugation were performed before or after freezing.

Orientadores: Maria Conceição da Costa, Rafael de Brito Dias / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Geociências / Made available in DSpace on 2018-08-21T19:16:42Z (GMT). No. of bitstreams: 1 Vicente_AlexandreMeloni_M.pdf: 2230181 bytes, checksum: ba205559be02d6ac3b4e95bacd04c731 (MD5) Previous issue date: 2012 / Resumo: A disseminação, principalmente a partir da década de 1970 nos países desenvolvidos do ocidente, de procedimentos participativos em diversas áreas, pautados na ideia de amplo engajamento dos cidadãos nos assuntos de interesse da coletividade, e em movimentos de pluralismo e democracia direta minou o modelo tradicional de governança onde os políticos eleitos, com a ajuda de experts reconhecidos, decidem as ações políticas sem maior interferência do público. O lugar da ciência na sociedade se torna cada vez mais problemático. A confiança pública no progresso da ciência e da tecnologia decresce, enquanto as preocupações sobre suas consequências crescem. A própria natureza da sociedade democrática demanda, deste modo, que a Ciência e sua influência estejam sujeitos a rigorosos processos de diálogo e crítica, e as controvérsias em ciência e tecnologia são cada vez mais reconhecidas como questões éticas e morais, e não somente técnicas. É o caso da experimentação animal, tema central deste trabalho. A partir do referencial teórico dos Estudos Sociais da Ciência e Tecnologia, em especial das noções de core set e participação pública, é feita uma análise da legislação do Estado de São Paulo sobre a experimentação animal, visando clarificar quais grupos tiveram seus interesses e reivindicações atendidos, e quais foram ignorados. O estudo serve de base para as considerações teóricas sobre o core set da controvérsia, abordando aspectos como a relação entre a comunidade científica, a camada política e o público leigo, a autoridade da ciência e as estratégias para inclusão/exclusão de grupos. Foi possível concluir que, apesar da diminuição da confiança pública no progresso da ciência e tecnologia, do enfraquecimento da credibilidade da opinião dos experts cientistas e dos crescentes questionamentos sobre as consequências físicas, sociais, éticas e morais da prática científica, a ciência ainda desempenha um papel central na resolução de controvérsias, e a comunidade científica ocupa um lugar privilegiado dentro do core set. Embora o diálogo com membros da sociedade civil seja necessário, a credibilidade e o status social privilegiado dos experts os possibilitam moldar o núcleo decisório de acordo com seus interesses, excluindo os grupos contrários à experimentação animal, e incluindo aqueles de ideologia moderada; mantendo, deste modo, um estável controle sobre a agenda de pesquisa / Abstract: The dissemination, mainly from the 1970s in Western developed countries, of participatory procedures in several areas, guided by the idea of broad citizen engagement on issues of interest to the community, and movements of direct democracy and pluralism, has undermined the traditional governance model where elected politicians, with the help of recognized experts, decide the political actions without interference from the public. The place of science in society becomes increasingly problematic. Public confidence in the progress of science and technology decreases as concerns grow about its consequences. The very nature of a democratic society demands thus that science and its influence are subject to rigorous review and dialogue processes, and controversies in science and technology are increasingly recognized as ethical and moral issues, not just technical. It is the case of animal experimentation, the central issue of this work. Using the theoretical framework of Social Studies of Science and Technology, particularly the concepts of core set and public participation, the work analyses the law of the State of São Paulo, Brazil, about animal experimentation, aiming to clarify which groups had their interests and demands met, and which were ignored. The study serves as basis for theoretical considerations on the core set of the controversy, addressing issues such as the relationship between the scientific community, the lay public and the policy layer, the authority of science and the strategies for inclusion/exclusion of groups. It was possible to conclude that, despite the decline of public confidence in the progress of science and technology, the weakening in the credibility of the expert's opinions, and the growing doubts about the physical, social, moral and ethical consequences of scientific practice, science still plays a central role in the resolution of controversies, and the scientific community occupies a privileged place within the core set. Although the dialogue with members of the civil society is necessary, the expert's credibility and privileged social status allows the scientific community to shape the core set according to its interests, excluding groups opposed to animal experimentation, and including those of moderate ideology; keeping thus a stable control over the research agenda / Mestrado / Politica Cientifica e Tecnologica / Mestre em Política Científica e Tecnológica

Orientador: Laymert Garcia dos Santos / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Filosofia e Ciências Humanas / Made available in DSpace on 2018-08-20T08:48:47Z (GMT). No. of bitstreams: 1 Santos_AndersonMarcosdos_D.pdf: 1864131 bytes, checksum: e679cd1dc91c477671aad93c467e5459 (MD5) Previous issue date: 2012 / Resumo: A tese discute os efeitos sociais da regulação jurídica das patentes sobre elementos biológico-informacionais humanos sob três aspectos: alteração do papel social do direito; a decisão política a respeito das opções tecnológicas; o delineamento de uma nova concepção de humano. Partimos da análise da aceleração como experiência central de temporalidade e como fenômeno transversal que perpassa o projeto moderno e o processo de modernização. Aceleração que atinge um ponto crítico com a virada cibernética em razão da guinada que esta provoca na lógica operatória da técnica ao possibilitar o acesso total ao controle instrumental pela reciprocidade informacional que permeia a matéria inerte, o ser vivo e o objeto técnico; e ao propiciar um rearanjo na aliança estabelecida entre o capital, a ciência e a tecnologia, que coloca a tecnociência como o motor de uma acumulação com a pretensão de abarcar todo o mundo existente, inclusive o humano, como matéria-prima à disposição do trabalho tecnoeconômico. Tendo como pressuposto a aceleração tecnoeconômica, percorremos a construção histórica do sistema internacional de patentes e os rearranjos nos requisitos de patenteabilidade ocorridos para adequá-los ao ritmo e à demanda dessa aceleração. Discutimos a formulação do discurso e da prática jurídicos para enquadrar os elementos biológico-informacionais humanos no sistema de patentes para, então, problematizarmos os riscos para o devir tecnológico, operado pelo bloqueio da modulação e da recombinação da informação, bem como a reconfiguração da concepção do humano que o direito está contribuindo para construir. Por fim, analisamos como o direito opera politicamente, mesmo que fora dos parâmetros do poder soberano clássico, quando assume uma posição diante da relação humano-máquina, reduzindo essa relação aos ditames da aceleração tecnoeconômica; e como o direito está mudando seu papel social, ao abandonar sua pretensão normatizadora e assumir um papel de regulador das relações sociais / Abstract: This dissertation discusses the social effects of the legal regulation of patents on human biologico-informational elements in view of three aspects: the change in the social role of Law; the political decision regarding technological options; the delineation of a new concept of human. It starts with the analysis of acceleration as a central temporality experience and as a transversal phenomenon that passes through the modern project and the modernization process. Acceleration reaches its critical point with the cybernetic upturn, derived from the shift that it provokes in the operating logic of technique as it allows full access to instrumental control thanks to the informational reciprocity that suffuses inert matter, living beings, and the technical object; and as it allows rearrangement in the alliance established among capital, science, and technology that places technoscience as the driver of kind of accumulation that intends to seize the whole existing world, including the human, as raw-material available to techno-economic labor. Premised on techno-economic acceleration, this paper goes over the historical construction of the international patent system and the rearrangements in patentability requisites made to adjust them to the rhythm and demand of such acceleration. It also discusses the formulation of legal discourse and practice to fit the human biologic-informational elements into the patent system, before problematizing the risks to technological upcoming developments, operated by the blockage of modulation and information recombination, as well as the reconfiguration of the concept of human that Law is helping to build. Lastly, the paper analyses how Law operates politically, even when outside the parameters of classic sovereign power, as it takes on a stance before the human-machine relationship, reducing this relationship to the dictates of techno-economic acceleration; and how Law is changing its social role by abandoning its standardizing intent to assume the role of social relationships regulator / Doutorado / Sociologia / Doutor em Sociologia

CYP2D6, a member of the cytochrome P450 enzyme system, has a central role in drug metabolism, it metabolizes 25 % of clinically used drugs. The gene that codes for the enzyme displays a high degree of polymorphism, which effects enzyme functions to various degrees. Aside from smaller mutations like SNPs, alleles may also feature duplications or deletion of the whole gene. Due to the clinical relevance of these mutations, a simple and precise method for genotyping is needed. In this study, a method based on pyrosequencing for copy number analysis was evaluated, wherein the copy number was determined by relative quantification to a reference gene CYP2D8P. During evaluation of the method, several adjustments were tried for optimization, including adjustments of annealing temperature and primer concentration. The results showed a difficulty in distinguishing between copy numbers using the method, as well as a high coefficient of variation. Therefore, further optimization is required before the method could be implemented into clinical practice.

Malignant lymphomas are a group of cancerous diseases that develop from lymphocytes and primarily affect lymph nodes. Being the sixth most common cancer type in Sweden, lymphoma is a societal problem that needs to be tackled by improving care and treatment of patients. This study was designed to examine the blood cell composition in lymphoma patients and well as determine whether the use of cryopreserved cells affected the analysis outcome. An evaluation of the methods used was also performed. Frozen peripheral blood from lymphoma patients as well as fresh and frozen blood from healthy controls was used. The cells of interest were monocytes, granulocytes, Treg, NKT, iNKT, B and T cells plus the dendritic cell activation protein CCR7. Three immunophenotyping methods were used. Method one was used in staining surface cell markers while the other two were for both surface and intracellular staining using two distinctive kits. The results showed no significant difference in immune cell composition between patients and blood donors. Limited patient samples and the lack of female blood donors could explain the unexpected result. A substantial difference in Treg cells was observed in fresh and frozen tested samples as well as T cell outcomes in method one compared to the other two methods. There were fewer Treg cells in frozen samples, which probably was due to cryopreservation while the lack of fixation in method one led to the loss of CD4+ T cells. Overall, the methods used were adequate but definitely require some improvements.

Escherichia coli are bacteria found in bowels of warm blooded animals. Most subspecies are harmless and part of the normal gut flora. However, E. coli have the ability to exchange genetic material with other bacteria, and some E. coli have acquired genes coding for virulence factors. VTEC, E. coli with the ability to produce verotoxin are commonly found in cattle, but certain types can cause severe disease in humans, known as enterohaemorrhagic E. coli, EHEC.     In this study, isolates of E. coli and other bacteria in the family Enterobacteriaceae from calves were subtyped and clustered using MALDI-TOF. Ten strains were selected for experimental competition analysis against E. coli MG1655.     The aim of the study was to identify strains of bacteria with the potential to outcompete VTEC in the cattle host and decrease the risk of human infections.     Three of the bacterial strains were able to outcompete the laboratory strain, and in future studies these strains can be analysed when competing against VTEC. The rest of the strains were outcompeted. Four known strains of VTEC were analysed competing the laboratory strain, showing weak ability to compete. Finally, a highly pathogenic strain of VTEC was analysed against Escherichia coli Nissle 1917, known for its ability to outcompete many strains of bacteria. Nissle could not outcompete the tested VTEC strain under the tested conditions.     In conclusion the majority of the bacterial strains isolated from calves were identified as E. coli and three of the isolates showed good ability to compete against the laboratory strain.