Made available in DSpace on 2016-04-28T20:15:33Z (GMT). No. of bitstreams: 1 2009 - Aline Rodrigues Bernardo.pdf: 1109499 bytes, checksum: 4b0f2adb9ceeb43406ffa4f7d20978ba (MD5) Previous issue date: 2009-02-09 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The production of specific antibody (IgY) in chickens (Gallus gallus domesticus) and its purification from the yolk of the egg is attracting the interest of the scientific community due to the numerous advantages of the IgG of mammals. Low cost, high volume production, recognition proteins highly conserved in mammals, in addition to preventing the suffering and death of animals for experimentation, are the main advantages. The aim of our study was to develop, standardize and deploy the technology IgY for aviaries production of antibodies with aim to obtain IGY-specific forms to promastigotes of Leishmania (Leishmania) amazonensis. Initially, matching was performed by two methods of precipitation and the comparison between them, to obtain the pure IgY from the yolk of egg white obtained in trade. The method of purification employing polietienoglicol (PEG-6000) was able to extract IgY with purity comparable to commercial standard IgY (Sigma) and more than obtained by the method with 25% ammonium sulfate. Later, chickens were inoculated with three doses of inactivated antigen (promastigotes forms of L. amazonensis) mixed with incomplete Freund's adjuvant, at 0, 14 and 21 days, IM route. High yield of total and specific IgY was obtained, as determined by the methods of Bradford and ELISA, respectively. The IgY antibodies showed high specificity (indirect immunofluorescence), sensitivity and avidity (ELISA) for promastigotes forms of L. amazonensis, from the second immunization. The level of IgY-specific antibodies remained high until the end of the analysis, around 56 days. Based on the results we can conclude that the "Technology IgY" allows to obtain antibodies with high specificity and sensitivity, in ethics, can be used successfully in various fields of science. / A produ??o de anticorpo espec?fico (IgY) em galinhas (Gallus gallus domesticus) e sua purifica??o a partir da gema do ovo vem atraindo o interesse da comunidade cient?fica devido as in?meras vantagens sobre os anticorpos IgG de mam?feros. Baixo custo, produ??o em quantidades elevada, reconhecimento de prote?nas altamente conservadas de mam?feros, al?m de se evitar o sofrimento e morte de animais de experimenta??o, s?o as principais vantagens. O objetivo geral de nosso estudo foi desenvolver, padronizar e implantar a Tecnologia IgY para produ??o de anticorpos avi?rios, tendo como meta a obten??o de IgY-espec?fica para formas promastigotas de Leishmania (Leishmania) amazonensis. Inicialmente, foi realizada adequa??o de dois m?todos de precipita??o e a compara??o entre eles, visando a obten??o da IgY pura a partir da gema de ovos brancos adquiridos no com?rcio. O m?todo de purifica??o empregando polietienoglicol (PEG-6000) foi capaz de extrair IgY com pureza compar?vel ao padr?o IgY comercial (Sigma) e superior a obtida pelo m?todo com sulfato de am?nio 25%. Posteriormente, galinhas foram inoculadas com tr?s doses de ant?geno inativado (formas promastigotas de L. amazonensis) misturados com adjuvante incompleto de Freund, nos tempos 0, 14 e 21 dias, por via intramuscular. Elevado rendimento da IgY total e espec?fica foi obtido, sendo determinado atrav?s dos m?todos de Bradford e ELISA, respectivamente. Os anticorpos IgY apresentaram elevada especificidade (Imunofluoresc?ncia indireta), sensibilidade e avidez (ELISA) para formas promastigotas de L. amazonensis, a partir da segunda imuniza??o. O n?vel de anticorpos IgY-espec?ficos permaneceu elevado at? o final da an?lise, por volta do 56? dia. Com base nos resultados obtidos podemos concluir que a Tecnologia IgY possibilita a obten??o de anticorpos com alta especificidade e sensibilidade, de forma ?tica, podendo ser utilizada com sucesso em diversos campo da ci?ncia.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Nos Ãltimos vinte anos, o campo de biomateriais à base de fosfato de cÃlcio tem crescido rapidamente, desempenhando um papel-chave como substituinte Ãsseo. A hidroxiapatita (HA) apresenta semelhanÃa quÃmica e estrutural com a fase mineral dos ossos e dos dentes, à biocompatÃvel e osteocondutiva, tem excelente afinidade quÃmica e biolÃgica com os tecidos Ãsseos e tem aplicaÃÃes biomÃdicas importantes como preenchimento de defeitos Ãsseos e em âscaffoldsâ na engenharia de tecidos. Nanocristais de HA foram sintetizados com sucesso por co-precipitaÃÃo e via hidrotÃrmica. Para a caracterizaÃÃo estrutural, tamanho e morfologia das fases obtidas foram utilizadas tÃcnicas de DifraÃÃo de Raios-x (DRX), Espectroscopia na RegiÃo do Infravermelho com Transformada de Fourier (FTIR), AdsorÃÃo/dessorÃÃo de N2, Potencial Zeta (PZ) e Microscopia EletrÃnica de TransmissÃo (TEM). Estruturas mesoporosas foram obtidas com diÃmetros de poros variando entre 12 e 31 nm. Os nanocristais de HA apresentaram uma morfologia definida, em forma de bastÃo, e foram incorporadas a um polÃmero para formaÃÃo de um âscaffoldâ. A droga sinvastatina (SINV), que influencia a regeneraÃÃo Ãssea quando aplicada localmente, foi associada aos biocompÃsitos obtidos. A caracterizaÃÃo dos âscaffoldsâ foi realizada por Microscopia EletrÃnica de Varredura (MEV). / In the last twenty years, the field of biomaterials based on calcium phosphates has grown rapidly, playing a key role as a bone substituent. Hydroxyapatite has chemical and structural similarity to the mineral phase of bone and teeth, is biocompatible and osteoconductive. It has excellent chemical and biological affinity with the bone tissue and important biomedical applications such as filling bone defects and scaffolds in tissues engineering. Hydroxyapatite nanocrystals were successfully synthesized by co-precipitation and hydrothermal route. X-ray diffraction, Fourier-transform infrared spectroscopy, N2 adsorption/desorption, Zeta Potential and Transmission Electron Microscopy were used to structural characterization, size and morphology of the nanoparticles. Mesoporous structures were obtained with pore diameters ranging from 12 to 31 nm. The nanoparticles showed a rod-shaped mophorlogy and were incorporated into a polymer to form a scaffold. The drug simvastatin, which influences bone regeneration when locally applied, was associated with the biocomposite. The characterization of the scaffolds was performed by Scanning Electron Microscopy.

<p>Cilj ove doktorske disertacije predstavljalo je ispitivanje uticaja vodnog stresa izazvanog&nbsp; sa polietilen glikolom (PEG)&nbsp; 6000 na biohemijske osobine klonova topole. Reznice tri klona topole (M-1, B-229 i PE19/66) su hidroponski gajene i izložene 100 mOsm i 200 mOsm PEG 6000&nbsp; tokom &scaron;est dana. Nakon tretmana u kontrolnim i stresiranim<br />biljkama su&nbsp; ispitane promene aktivnosti antioksidantnih enzima,&nbsp; aktivnost dva<br />enzimska markera polifenolnog metabolizma, fenolni profil, antioksidantna aktivnost, kao i sadržaj prolina i glicin betaina&nbsp; (GB). Takođe, ispitan je biohemijski odgovor kulture tkiva klona M-1 na vodni stres izazvan sa PEG 6000.</p><p>U oba eksperimenta uočeno je da je akumulacija prolina i GB u uslovima vodnog stresa izazvanog sa PEG 6000&nbsp;najvažnija strategija u otpornosti na stres i prevazilaženju vodnog deficita, tako da se&nbsp;ovi parametri mogu definisati kao najbolji indikatori otpornosti topole na su&scaron;u.</p><p>Rezultati ovog istraživanjaupotpunjavaju saznanja o povezanosti oksidativnog stresa uzrokovanog vodnim stresom sa antioksidantnim odgovorom, nivoom o&scaron;tećenja lipida i proteina, uključivanjem osmolita kao i promenama u polifenolnom profilu. Takođe, dobijeni rezultati su značajni za odabir klonova topola tolerantnih na vodni stres&nbsp; i&nbsp; za<br />po&scaron;umljavanje su&scaron;nih stani&scaron;ta, kojih će biti sve vi&scaron;e usled negativnog uticaja klimatskih promena.</p> / <p>The aim of presented doctoral thesis was investigation of the impact of water deficit caused by polyethylene glycol (PEG) 6000 on biochemical features of poplar clones.&nbsp; During six days, cuttings of three poplar clones (M-1, B-229 and PE 19/66) were grown in hydroponics and subjected to 100 and 200 mOsm PEG 6000. Changes in the activity of antioxidant enzymes, polyphenol characterization, two enzymatic markers of polyphenol metabolism, antioxidant capacity, as well as proline and glycine betaine content were investigated in stressed plants. Furthermore,&nbsp; assessment of&nbsp; biochemical response of poplar clone M-1 tissue culture to water stress induced by PEG 6000 was performed.</p><p>In both experiments, the most important strategy for stress resistance and overcoming water deficiency was accumulation of proline and glycine betaine so these parameters can be defined as the best indicators of poplar resistance to drought.</p><p>The results of this study complete&nbsp; the findings on the oxidative stress caused by water stress&nbsp; and their relationship with the antioxidant response, the level of lipid andprotein damage, accumulation of the osmolites and changes in the polyphenol profile. Also, the obtained results may be important for the selection of poplar clones resistant to water stress and for afforestation in arid sites, which appeared to be more numerous due to the effects of&nbsp; global climate change.</p>