Aseptic loosening of the components is probably the most common long-term complication resulting in failure of Total Hip Arthroplasty. The mechanical behaviour of bone under load is one of the contributory causes of loosening encountered at the prosthesis/cement/bone interface. The present study dealt with a series of invitro experiments conducted on epoxy resin models of human hemi-pelves with different commercially available acetabular components implanted in them. These are used for the construction of simplified models of the artificial hip joint (three-dimensional) and of the prosthesis/cement/bone acetabular interface (two-dimensional). Loading conditions for the models included tensioning of the simulated abductor muscles for the hemi-pelvic and femoral loading for the prosthesis/cement/bone interface study. The experimental method employed was real-time holographic interferometry, a stress analysis technique recently used in the biomechanical field, which permitted whole-field simultaneously inspection of deformation patterns. The holographic interferograms were interpreted in a qualitative rather than a quantitative manner. The models do not exactly represent the in-vivo situation. Since this study identified high stresses both in the hip bone as well as in the interface (prosthesis/bone) it is suggested that these stresses are implicated in the mechanical pathogenesis of loosening. The observed changes in stress levels detected in our models could serve as a guide for future designs of acetabular prostheses as well as guide a in surgical techniques.

Xanthene dyes are common fluorophores which have been widely used as molecular probes. The xanthene fluorophores can be used as highly selective optical sensors to detect disease biomarkers. A new fluorogenic dye containing an alpha, beta-unsaturated aldehyde moiety exhibits selective fluorescent signal enhancement in the presence of cysteine or peptides containing N-terminal cysteine residues. The mechanism is based on synergistic covalent and supramolecular interactions. A unique rhodamine boronic acid indicator is used in an optimized data collection protocol for wavelength- and time-dependent selectivity of various saccharides and nucleosides. One indicator is thereby capable of selectively distinguishing structurally related analytes in mixtures. Moreover, the rhodamine-based boronic acid responds linearly to increasing riboside concentrations in urine samples, potentially enabling the screening for inborn purine metabolism disorders.

Na realização da colangiopancreatografia endoscópica retrógrada a cateterização da papila duodenal maior é passo fundamental na obtenção do acesso biliar profundo e correlaciona -se com complicações biliopancreáticas das quais a pancreatite aguda pós-CPER é a mais comum. Os objetivos deste trabalho foram: a) comparar o índice de sucesso na canulação seletiva da via biliar com uso do canulótomo e canulótomo com fio guia; b) comparar, entre ambos os grupos, as dosagens séricas de amilase, lipase e proteína C reativa; c) avaliar a incidência de pancreatite nos grupos em estudo. No período de julho de 2002 a outubro de 2003 foram realizadas 341 CPER em três Instituições de nível terciário, destas foram randomizados prospectivamente e de maneira consecutiva 300 pacientes para cateterização papilar com canulótomo (Grupo I) e canulótomo com fio guia (Grupo II). Os procedimentos endoscópicos foram realizados pelo autor nas três Instituições. Procedeu-se a caracterização do perfil técnico-laboratorial e avaliação da incidência de pancreatite através de métodos clínicolaboratoriais e imagenológicos, para ambos os grupos. Todos os pacientes do estudo foram mantidos internados por 24 horas após a CPRE. A cateterização inadvertida do ducto pancreático foi semelhante para os dois grupos (p= 0,161). A fistulopapilotomia foi mais freqüente no grupo I (p= 0,011), porém apresentou significativamente menor incidência de pancreatite aguda no grupo II (p= 0,041). As dosagens séricas de amilase coletadas quatro, 12 e 24 horas após CPER foram significativamente maior no grupo I (p= 0,0087; p= 0,045; p= 0,0474; respectivamente). As dosagens séricas de lipase e proteína C-reativa após a CPER foram similares para ambos os grupos. O tempo de manipulação pancreática apresentou elevação similar nas dosagens séricas de amilase após a CPRE, porém todas as dosagens de lipase coletadas após a CPER foram significativamente maior no grupo I para a categorização de um a cinco minutos (p= 0,025; p= 0,032; p= 0,049). O número de cateterizações pancreáticas categorizadas em uma a cinco vezes apresentou elevação significativamente maior no grupo I, para as amostras de amilase, lipase e proteína C-reativa coletadas quatro, 12 e 24 horas após a CPER (amilase: p=0,006; p= 0,0023; p= 0,0095/lipase: p= 0,13; p= 0,018; p= 0,028 / PC-R: p= 0,005; p= 0,01; p= 0,01). As papilotomias realizadas no grupo II apresentaram significativamente maior elevação das dosagens séricas de amilase coletadas 12 e 24 horas após a CPER (p= 0,033; p= 0,049). As dosagens séricas de lipase e proteína C-reativa apresentaram elevações similares tanto na papilotomia como na fistulopapilotomia. A pancreatite aguda pós-CPER foi significativamente maior no grupo I (p= 0,037). Conclusões: a) O acesso biliar através do cateter com fio guia proporcionou maior índice de sucesso na canulação biliar seletiva; b) No perfil laboratorial estudado a dosagem de amilase se mostrou com diferença significante na comparação entre os grupos estudados. O mesmo não ocorreu nas dosagens de lipase e PC-R; c) O uso do fio guia foi um fator de prevenção na ocorrência da pancreatite aguda pós-CPRE / During the endoscopic retrograde cholangiopancreatography (ERCP) the main step is the cannulation of major duodenal papilla to obtain deep bile duct access, and it is correlated to pancreaticobiliary complications being acute pancreatitis the most frequent. The aims were: a) compare the rate of success to achieve selective cannulation of common bile duct using a single cannula and cannula with guide-wire; b) compare the amylase, lypase and Creactive protein serum level between the groups; c) evaluate the incidence of pancreatitis in the groups. From July 2002 to October 2003 there were performed 341 ERCP on three institutions of tertiary level. From them, 300 patients were randomized, on a prospective and consecutive fashion to major duodenal papilla cannulation using single cannula (Group I) and cannula with guide wire (Group II). The author himself performed all the endoscopic procedures on the three institutions. The characterization of technicallaboratory profile and evaluation of the incidence of pancreatitis were proceeded by clinical-laboratory and image methods to both groups. All patients were hospitalized by 24 hours after ERCP. The cannulations of pancreatic duct were similar to both groups (p=0,161). The fistulosphincterotomy was more frequent in group I (p=0,011), but group II presented significant lower incidence of acute pancreatitis (p=0,041). The amylase serum were collected 4, 12 and 24 hours after ERCP and were significantly higher in group I (p=0,0087; p=0,045; p=0.0474, respectively). The lypase and C-reactive protein after ERCP were similar to both groups. The time of pancreatic manipulation presented similar elevation of amylase serum after ERCP, therefore all lypase serum after ERCP were significantly higher in group I for the categorization of 1 to 5 minutes (p=0,025; p=0,032;p=0,049). The number of pancreatic cannulations categorized in 1 to 5 times presented significant higher elevation in group I, to the samples of amylase, lypase and C-reactive protein serum collected 4, 12 and 24 hours after ERCP (amylase: p=0,006; p=0,0023; p=0,0095/ lypase: p=0,13; p=0,018;p=0,028/ C-RP: p=0,005; p=0,01; p=0,01). The endoscopic papillotomy performed in group II presented significant higher elevation of amylase serum collected at 12 and 24 hour post ERCP (p=0,033;p=0,049). The lypase and C-reactive protein serum presented similar elevation such as in papillotomy as in fistulosphincterotomy. The acute pancreatitis post ERCP were significantly higher in group I (p=0,037). Conclusion: a) The biliar access by cannula with guide wire offered a higher success to selective biliar cannulation; b) the laboratory profile of amylase serum showed a significant difference between the groups. It did not occur with lypase and C-reactive protein serum levels; c) the use of guide wire was a preventing factor of acute pancreatitis post ERCP

Na realização da colangiopancreatografia endoscópica retrógrada a cateterização da papila duodenal maior é passo fundamental na obtenção do acesso biliar profundo e correlaciona -se com complicações biliopancreáticas das quais a pancreatite aguda pós-CPER é a mais comum. Os objetivos deste trabalho foram: a) comparar o índice de sucesso na canulação seletiva da via biliar com uso do canulótomo e canulótomo com fio guia; b) comparar, entre ambos os grupos, as dosagens séricas de amilase, lipase e proteína C reativa; c) avaliar a incidência de pancreatite nos grupos em estudo. No período de julho de 2002 a outubro de 2003 foram realizadas 341 CPER em três Instituições de nível terciário, destas foram randomizados prospectivamente e de maneira consecutiva 300 pacientes para cateterização papilar com canulótomo (Grupo I) e canulótomo com fio guia (Grupo II). Os procedimentos endoscópicos foram realizados pelo autor nas três Instituições. Procedeu-se a caracterização do perfil técnico-laboratorial e avaliação da incidência de pancreatite através de métodos clínicolaboratoriais e imagenológicos, para ambos os grupos. Todos os pacientes do estudo foram mantidos internados por 24 horas após a CPRE. A cateterização inadvertida do ducto pancreático foi semelhante para os dois grupos (p= 0,161). A fistulopapilotomia foi mais freqüente no grupo I (p= 0,011), porém apresentou significativamente menor incidência de pancreatite aguda no grupo II (p= 0,041). As dosagens séricas de amilase coletadas quatro, 12 e 24 horas após CPER foram significativamente maior no grupo I (p= 0,0087; p= 0,045; p= 0,0474; respectivamente). As dosagens séricas de lipase e proteína C-reativa após a CPER foram similares para ambos os grupos. O tempo de manipulação pancreática apresentou elevação similar nas dosagens séricas de amilase após a CPRE, porém todas as dosagens de lipase coletadas após a CPER foram significativamente maior no grupo I para a categorização de um a cinco minutos (p= 0,025; p= 0,032; p= 0,049). O número de cateterizações pancreáticas categorizadas em uma a cinco vezes apresentou elevação significativamente maior no grupo I, para as amostras de amilase, lipase e proteína C-reativa coletadas quatro, 12 e 24 horas após a CPER (amilase: p=0,006; p= 0,0023; p= 0,0095/lipase: p= 0,13; p= 0,018; p= 0,028 / PC-R: p= 0,005; p= 0,01; p= 0,01). As papilotomias realizadas no grupo II apresentaram significativamente maior elevação das dosagens séricas de amilase coletadas 12 e 24 horas após a CPER (p= 0,033; p= 0,049). As dosagens séricas de lipase e proteína C-reativa apresentaram elevações similares tanto na papilotomia como na fistulopapilotomia. A pancreatite aguda pós-CPER foi significativamente maior no grupo I (p= 0,037). Conclusões: a) O acesso biliar através do cateter com fio guia proporcionou maior índice de sucesso na canulação biliar seletiva; b) No perfil laboratorial estudado a dosagem de amilase se mostrou com diferença significante na comparação entre os grupos estudados. O mesmo não ocorreu nas dosagens de lipase e PC-R; c) O uso do fio guia foi um fator de prevenção na ocorrência da pancreatite aguda pós-CPRE / During the endoscopic retrograde cholangiopancreatography (ERCP) the main step is the cannulation of major duodenal papilla to obtain deep bile duct access, and it is correlated to pancreaticobiliary complications being acute pancreatitis the most frequent. The aims were: a) compare the rate of success to achieve selective cannulation of common bile duct using a single cannula and cannula with guide-wire; b) compare the amylase, lypase and Creactive protein serum level between the groups; c) evaluate the incidence of pancreatitis in the groups. From July 2002 to October 2003 there were performed 341 ERCP on three institutions of tertiary level. From them, 300 patients were randomized, on a prospective and consecutive fashion to major duodenal papilla cannulation using single cannula (Group I) and cannula with guide wire (Group II). The author himself performed all the endoscopic procedures on the three institutions. The characterization of technicallaboratory profile and evaluation of the incidence of pancreatitis were proceeded by clinical-laboratory and image methods to both groups. All patients were hospitalized by 24 hours after ERCP. The cannulations of pancreatic duct were similar to both groups (p=0,161). The fistulosphincterotomy was more frequent in group I (p=0,011), but group II presented significant lower incidence of acute pancreatitis (p=0,041). The amylase serum were collected 4, 12 and 24 hours after ERCP and were significantly higher in group I (p=0,0087; p=0,045; p=0.0474, respectively). The lypase and C-reactive protein after ERCP were similar to both groups. The time of pancreatic manipulation presented similar elevation of amylase serum after ERCP, therefore all lypase serum after ERCP were significantly higher in group I for the categorization of 1 to 5 minutes (p=0,025; p=0,032;p=0,049). The number of pancreatic cannulations categorized in 1 to 5 times presented significant higher elevation in group I, to the samples of amylase, lypase and C-reactive protein serum collected 4, 12 and 24 hours after ERCP (amylase: p=0,006; p=0,0023; p=0,0095/ lypase: p=0,13; p=0,018;p=0,028/ C-RP: p=0,005; p=0,01; p=0,01). The endoscopic papillotomy performed in group II presented significant higher elevation of amylase serum collected at 12 and 24 hour post ERCP (p=0,033;p=0,049). The lypase and C-reactive protein serum presented similar elevation such as in papillotomy as in fistulosphincterotomy. The acute pancreatitis post ERCP were significantly higher in group I (p=0,037). Conclusion: a) The biliar access by cannula with guide wire offered a higher success to selective biliar cannulation; b) the laboratory profile of amylase serum showed a significant difference between the groups. It did not occur with lypase and C-reactive protein serum levels; c) the use of guide wire was a preventing factor of acute pancreatitis post ERCP

Thesis (MScMed)--Stellenbosch University, 2008. / ENGLISH ABSTRACT: Introduction: Flow cytometry has progressively replaced many traditional laboratory tests due to its greater accuracy, sensitivity and rapidity in the routine clinical settings especially clinical trails. It is a powerful tool for the measuring of chemical (the fluorochrome we add) and physical (size and complexity) characteristics of individual cells. As these instruments became major diagnostic and prognostic tools, the need for more advanced quality control, standardized procedures and proficiency testing programs increased as these instrumentations and their methodology evolve. Minor instrument settings can affect the reliability, reproducibility and sensitivity of the cytometer and should be monitored and documented in order to ensure identical conditions of measurement on a daily basis. This can be accomplished by following an Internal Quality Assurance (IQA) and/ or External Quality Assurance (EQA) program. Currently there are no such programs available in South Africa and poorer Africa countries. HIV is a global concern and the laboratories and clinics in these places are in need of such IQA programs to ensure quality of their instrumentation and accurate patient results. Quality assurance programs such as CD Chex® and UK Nequas are available but due to bad sample transport, leave the receiving laboratories with nightmares. It would be best if there was a laboratory in South Africa that could provide the surrounding laboratories with stabilized whole blood samples that can be utilized as IQA. The transport of these samples can be more efficient due to shorter distance and thus the temperature variations limited. Aims and Objectives: The aim of Chapter one is to familiarize the reader with general terminology and concepts of immunology. Chapter two describes in detail the impact stabilized whole blood had on clinical immunology concerning Quality Control and Quality Assurance. The objective of this study is to stabilize whole blood with a shelf life of greater than 30 days to serve as reference control material for South African Immunophenotyping. It is further an objective to use these in-house stabilized control samples for poorer African countries as Internal Quality Assurance reference material. It is a still further objective to stimulate various lymphocyte subsets to express activation antigens and then stabilize these cells for more specialized immunological test and can serve as a QC for those required samples. Study design: In Chapter three, the method currently used to stabilize whole blood was modified. The stability of different concentrations of a first stabilizing agent (Chromium Chloride hexahydrate) was investigated. Incubation periods and concentrations of paraformaldehyde as second stabilizing agent were investigated. Blood samples from healthy individuals (n=10) were stabilized and monitored for the routine HIV phenotypic surface antigens over a period of 40 days. These samples (n=10) were compared on the Becton Dickinson Biosciences (BD) FACSCalibur™ versus BD FACSCount™ instrumentation. Blood samples (n=3) were stabilized and monitored to identify phenotypic cell surface molecules for as long as possible. They were quantified on both flow cytrometric instruments. In addition, these stabilized samples (n=3) were investigated as control blood for calibration purposes on the BD FACSCount™ instrument. In Chapter four, lymphocytes were isolated and activated with various stimuli to express sufficient activation antigens such as CD25, CD69, HLA-DR and CD40 Ligand on the T helper cell surfaces. These activated antigens were analyzed on the BD FACSCalibur™ and further stabilized to serve as possible IQA samples in future. Results: In Chapter three, the ten individual stabilized samples had non-significant P values (P > 0.05) for CD3, CD4 and CD8 percentages and absolute values comparing day 3 until day 40. Comparing the BD FACSCalibur™ versus BD FACSCount™, resulted in a R2 = 0.9848 for CD4 absolute values and a R2 = 0.9636 for CD8 absolute values. Stabilized blood samples (n=3) were monitored for routine HIV phenotypic markers until day 84. The cells populations were easily identifiable and could be quantified on both BD FACSCalibur™ and BD FACSCount™ instruments. In Chapter four; for the activation study purposes, activated T helper lymphocytes expressed approximately 25 to 35% CD40 Ligand cell surface molecules. The stimulant of choice was Ionomycin at a 4μM concentration. Cells were incubated for four hours at 37 degree Celsius in a 5% CO2 environment. For CD69 surface expression, 6 hour incubation was optimum. The stimulus of choice in this case was 4μM Ionomycin which induced 84.21% CD69 expression in the test samples. For CD25 expression; 6 hour incubation with PHA resulted in approximately 43% of CD25 expression. For HLA-DR surface expression; 6 hour incubation with PHA resulted in approximately 43.32% of HLA-DR expression. Activated lymphocytes expressing CD40 Ligand showed stability until day 23. Activated Lymphocytes expressing CD69, CD25 and HLA-DR were stabilized in the same manner and stability could be achieved until day 16. Conclusion: This thesis was related to the preparation of control samples (IQA) designed to simulate whole blood having defined properties in clinical laboratory situations. In future kits can be developed with a low, medium and high control sample for the various immunological phenotypic determinants. Another kit can be compiled where various activation markers can be identified, quantified with a “zero”, low and high control. These whole blood IQA kits and “activation IQA kits” can be implemented for training of newly qualified staff, competency testing of staff, method development, software testing, panel settings and instrument setting testing. Control samples ideally must have a number of properties in order to be effective. For instance stability during storage times, preferably lasting more than a few weeks, reproducibility and ease of handling. These will provide the information on day-to-day variation of the technique or equipment which will enhance accuracy and improve patient care. / AFRIKAANSE OPSOMMING: Inleiding: Vloeisitometrie tegnologie het verskeie tradisionele laboratorium toetse vervang as gevolg van beter akuraadheid, sensitiwiteit en vinniger beskikbaarheid van resultate in ‘n kliniese omgewing, veral kliniese proewe. Vloeisitometrie is ‘n kragtige tegniek om chemiese (fluorokroom byvoeging) en fisiese (sel grote en kompleksiteit) karakter eienskappe van individuele selle te meet. Met die toename in gebruik en gewildheid van hiedie instrumente, neem die behoefde toe vir gevorderde kwaliteit kontroles, gestandardiseerde prosedures, met profesionele toets programme tesame met metode ontwikkeling. Klein verstellings aan instrument parameters beinvloed die betroubaarheid, herhaalbaarheid en sensitiwiteit van ‘n sitometer en moet gemonitor (en dokumenteer) word om identiese kondisies van leesings op ‘n daaglikse basis te verseker. Dit kan bereik word deur in te skakel met ‘n interne kwaliteits versekerings program [IQA: “Internal Quality Control”] en/of ‘n eksterne kwaliteits versekerings program [EQA: “External Quality Control”] te volg. Op die oomblik is daar geen sulke kwaliteits versekerings programme in Suid Afrika en/of in die verarmende Afrika lande beskikbaar nie. MIV is ‘n wêreldwye bekommernis en laboratoriums en klinieke in hierdie gedeeltes van die land verlang ‘n dringende behoefdte vir sulke “IQA” programme om kwaliteit van instrumentasie en akkurate pasiënt resultate te verseker wat tot beter behandeling van pasiënte lei. Kwaliteit versekerings programme soos “CD Chex®” en “UK Nequas” is beskikbaar, maar baie probleme met verwysing na monster integriteit as gevolg van tydsame vervoer en aflewering kondisies word hiermee geassosieër. Die behoefte het ontstaan vir ‘n laboratorium in Suid Afrika wat direk die omliggende laboratoriums, hospitale en klinieke kan voorsien met gestabiliseerde blood monsters wat gebruik kan word as “IQA”. Die vervoer en aflewerings kondisies van hierdie monsters sal aansienlik verbeter as gevolg van die korter aflewerings afstand wat direk die beperkte temperatuur wisseling beinvloed. Doel van studie: Die doelwit van hoofstuk een is om vir die leser ‘n inleiding te gee tot terminologie en konsepte van immunologie en die immune sisteem. Hoofstuk twee beskyf die impak wat gestabiliseerde heelbloed het op die kliniese immunologie met betrekking tot kwaliteit beheer en kwaliteit versekering. Die doelwit van hierdie studie is om heelbloed te stabiliseer sodat die rakleeftyd meer as 30 dae is en sodoende as verwysings-materiaal kontroles vir Suid Afrikaanse immunofenotipering kan dien. Dit is ‘n verdere doelwit om hierdie tuis-gestabiliseerde kontrole monsters te gebruik as “IQA” verwysings materiaal in verarmende Afrika lande. Die doelwit van hoofstuk vier is om limfosiete te stimuleer om verskeie aktiverings merkers uit te druk op hul selmembrane en dan te stabiliseer en dié te gebruik as Kwaliteits Kontroles vir die meer gespesialiseerde immunologiese toetse. Studie ontwerp: Hoofstuk drie beskryf ‘n aangepaste en verbeterde metode van heel bloed stabiliseering. Stabiliteit word ondersoek in ‘n verskyndenheid konsentrasies van ‘n primêre stabiliseerings agent (chromium chloried heksahidraat) en inkubasie periodes met paraformaldehied as tweede stabiliseerings agent word deeglik gedokumenteer. Bloedmonsters van gesonde indiwidië (n=10) was gestabiliseer en gemonitor vir roetine MIV membraanoppervlak antigene oor ‘n periode van 40 dae. Hierdie monsters (n=10) was gelees en geanaliseer op ‘n BD FACSCalibur™ en vergelyk met ‘n BD FACSCount™ vloeisitometer instrument. Drie gestabiliseerde heelbloed monsters (n=3) was gemonitor vir ‘n periode vir so lank moontlik die fenotipiese selmembraan molekules identifiseerbaar was en die kwantiteit bepaalbaar was. Hierdie drie monsters was gemeet op beide instrumente. As ‘n addisionele doelwit, was hierdie drie gestabiliseerde monsters ondersoek om as moontlike kalibrasie materiaal (verteenwoordig ‘n normale bloedmonster) te dien vir die BD FACSCount™ instrument in die oggende voor pasiënt monsters gelees kan word. In hoofstuk vier was limfosiete geϊsoleer en geaktiveer met ‘n verskyndenheid stimulante om optimale aktiveerings-antigene uit te druk op T helper selmembrane (byvoorbeeld CD25, CD69, HLA-DR en CD40 Ligand). Hierdie geaktiveerde monsters was geanaliseer op die BD FACSCalibur™ en daarna gestabiliseer. Na stabilisasie van die geaktiveerde limfosiet monsters was dit gemonitor oor ‘n tydperk so lank moontlik data plotte leesbaar en selpopulasies identifiseerbaar was. Hierdie monsters kan dien as ‘n moontlike “IQA” toets stel vir ‘n meer gespesialiseerde immunologiese aktiveerings kontrole doeleindes. Resultate: In hoofstuk drie; tien individiële gestabiliseerde heelbloed monsters het gedui op geen-beduidende P waardes (P > 0.05) vir CD3, CD4 en CD8 persentasies en absolute waardes; gemeet vanaf DAG 3 vergelykbaar tot-en-met DAG 40. Met korrelasie statistiek en vergelyking van die BD FACSCalibur™ met die FACSCount™ instrumente, is die volgende opgemerk; R2 = 0.9848 vir die CD4 absolute waardes en ‘n R2 = 0.9636 vir die CD8 absolute waardes. Drie gestabiliseerde monsters (n=3) was gemonitor vir MIV roetine fenotipeering tot en met DAG 84. Die selpopulasies was duidelik identifiseerbaar en die kwantitatief meetbaar op albei instrumente (BD FACSCalibur™ en BD FACSCount™). Hoofstuk vier: geaktiveerde T helper lymphosiete het 25 – 35% membraan CD40 Ligand uitgedruk op hul selmembrane. Die stimulant van keuse was ionomysien teen ‘n optimale konsentrasie van 4μM. Die optimale inkubasie tydperk was vier ure by 37°C in 5% CO2 kondisie. Ses uur inkubasie in 4μM ionomysien by 37°C in ‘n 5% CO2 omgewing was optimal vir die CD69 selmembraan uitdrukking en het 84.21% opgelewer. Vir CD25 selmembraan uitdrukking was die selle vir ses ure met phietoheamagglutinin (PHA) gestimuleer by 37°C in 5% CO2 kondisie en het 43% CD25 selmembraan uitdrukking opgelewer. HLA-DR selmembraan uitdrukking: selle was vir ses ure saam met PHA by 37°C in 5% CO2 kondisie inkubeer en het 43.32% opgelewer. CD40 Ligand aktivering/gestabiliseerde limfosiete het tot en met dag 23 stabiliteit getoon. Die ligand was duidelik identifiseerbaar en kwantifiseerbaar. Geaktiveerde lymphosiete wat CD69, CD25 en HLA-DR selmembraan merkers uitdruk het na die stabiliseerings proses stabiliteit getoon tot-en-met dag 16. Gevolgtrekking: Die doel van hierdie studie was om verwysingskontroles voor te berei sodat dit vars heelbloed naboots met uitkenbare eienskappe vir kliniese situasies. ‘n Toets kontrolestel met verwysings materiaal vir drie vlakke (byvoorbeeld ‘n lae, medium en hoë kontrole) absolute selwaardes en persentasies kan voorberei word vir roetine immunologiese fenotiperings merkers (CD3/CD4/CD8/CD45). Meer gespesialiseerde kontrolestelle vir meer spesifieke doeleindes kan opgemaak word wat ‘n verskydenheid van limfosiet aktiveringsmerkers bevat met byvoorbeeld ‘n “nul”, lae en hoë verwysings kontrole daarin. Hierdie heelbloed kan dien as “aktiveerde interne kwaliteits verwysings materiaal” en kan gebruik word om nuut aangestelde laboratorium werkers en nuut gekwalifiseerde studente op te lei. Hierdie verwysings materiaal / kontroles kan aangewend word vir bevoegdheids doeleindes (byvoorbeeld vir SANAS akkreditasie doeleindes), vir metode ontwikkeling, vir sagteware toetsing, vir paneel opstelling en instrument verstellings doeleindes. Die kontroles moet ‘n verskydenheid eienskappe bevat om effektief te wees. Byvoorbeeld, stabiliteit tydens storing, gewenslik meer as ‘n paar weke, herhaalbaar en maklik handteerbaar. Hierdie kontroles sal inligting voorsien op ‘n daaglikse basis tydens wisseling van tegnieke of instrumentasie wat akuraatheid beinvloed en op die ou-end direk pasiënt versorging bevoordeel.

Thesis (MScMedSc)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Setting This study was conducted in the Tygerberg district, Cape Town, in the Western Cape, South Africa Background The evaluation of early tuberculosis (TB) treatment response is based on month 2 sputum culture status. This method of evaluation has a number of limitations: the test requires relatively advanced laboratory infrastructure and procedures, it takes several weeks to obtain results and is a relatively a poor marker at predicting treatment response. The discovery of potential host markers which reflect the efficacy of early treatment would be of great importance for clinical management of individual patients. The treatment failure would be detectable earlier than at week 8 of treatment. The duration of clinical trials of new anti-tuberculosis drugs may also be substantially reduced by such markers if these would be measurable earlier than at week 8 of therapy. Objectives 1) To evaluate diluted, 7-day whole blood cultures stimulated with live Mycobacterium tuberculosis (M.tb) for the presence of host markers of early TB treatment response 2) To evaluate an overnight, undiluted, M.tb antigen stimulated whole blood culture Quantiferon Gold In Tube (QFT-GIT) supernatants for host markers of early TB treatment response The study designs were as follows: In study one, baseline samples and samples from week 1, week 2 and week 4 of treatment from 30 cured TB patients were selected from a larger biomarker study, in which whole blood was stimulated with live M.tb or left unstimulated. Fifty seven host markers were measured in supernatants by multiplex cytokine arrays. In study two, baseline samples and samples from week 2 and week 8 of treatment from 19 cured TB patients were randomly selected from the placebo group in a micronutrient supplement study. QFT-GIT supernatants from these participants were assessed through multiplex cytokine arrays for levels of fifty seven host markers. All of the participants in both studies were Human Immunodeficiency Virus (HIV) negative. Changes in marker expression over time and between fast and slow responders to treatment were evaluated. Comparability between the two culture methods was assessed for markers that were evaluated in both studies. Results In study one, the majority of host markers showed significant changes over time in the unstimulated supernatants. Only GRO and IL-1beta changed significantly in an antigen-specific manner (background levels subtracted). No significant changes were observed between fast and slow responders. In study two, the majority of host markers showed significant changes over time in the unstimulated supernatants whereas only MDC and IL-4 changed during the observation period in antigen stimulated levels. Significant differences were observed between fast and slow responders at pre-treatment for IL-13 Ag-Nil and IL-1betaAg-Nil . Conclusion This study revealed, antigen-specific responses showed only limited potential for early TB treatment response monitoring, but may have potential in differentiating between treatment outcomes. Future investigations may have to include later time points during treatment as these were not included in the present assessment. The QFT-GIT samples do not appear to be equivalent to live M.tb stimulated 7-day whole blood assays. / AFRIKAANSE OPSOMMING: Instelling Die studie is uitgevoer in die Tygerbergdistrik, Kaapstad, Wes-Kaap, Suid-Afrika. Agtergrond Die evaluering van die respons op vroeë tuberkulose (TB) behandeling word gebaseer op die status van maand 2 sputum kulture. Hierdie evalueringsmetode het ‘n paar beperkinge: die toets benodig relatief gevorderde laboratorium infrastruktuur en prosedures, die toetsuitslae is eers na ‘n paar weke beskikbaar en dit is n relatiewe swak merker om repons op behandeling te voorspel. Die ontdekking van potensiële selfmerkers wat die doeltreffendheid van vroeë behandeling weerspieël sal van groot belang wees vir die kliniese bestuur van individuele pasiënte. Mislukking van die behandeling sal sodoende voor week 8 van behandeling waargeneem kan word. Die tydsduur van kliniese proewe van nuwe anti-tuberkulose medikasie mag ook baie verkort word met sulke merkers as dit voor week 8 van behandeling gemeet kan word. Doelwitte 1) Om verdunde, 7-dae oue volbloedkulture, met lewende Mikobakterium tuberkulosis (M.tb) gestimuleer, te evalueer vir die teenwoordigheid van vroeë TB behandeling respons selfmerkers. 2) Om die supernatant van oornag, onverdunde, M.tb antigeen gestimuleerde volbloedkulture Quantiferon Gold In Tube (QFT-GIT) vir vroeë behandeling respons selfmerkers te evalueer. Die studie-ontwerpe was soos volg: Met studie een is basislynmonsters en monsters verkry na week 1, week 2 en week 4 van behandeling van 30 geneesde TB-pasiënte geselekteer uit ‘n groter biomerkerstudie waarin die volbloed met lewende M.tb gestimuleer is of ongestimuleer gelaat is. Sewe-en-vyftig selfmerkers is in die supernatante gemeet deur middel van multipleks sitokine arrays. Met studie twee is basislynmonsters en monsters verkry na week 2 en week 8 van behandeling van 19 geneesde TB-pasiënte lukraak uit die plasebo-groep in ‘n mikrovoedingstowwe-aanvullingstudie geselekteer. Vlakke van 57 selfmerkers is in die QFT-GIT supernatante van hierdie deelnemers, deur middel van die multipleks sitokine arrays, bepaal. Al die deelnemers van beide studies was HIV negatief. Veranderinge in merker-uitdrukking oor tyd, asook tussen vinnige en stadige respons tot behandeling, is ge-evalueer. Die vergelykbaarheid van die twee kultuurmetodes is geassesseer ten opsigte van die ge-evalueerde merkers in albei studies. Resultate Met studie een het die meerderheid van die selfmerkers in die ongestimuleerde supernatante kenmerkende verandering oor tyd gewys. Slegs GRO en IL-1beta het aansienlik verander in die antigeenspesifieke wyse (agtergrond vlakke afgetrek). Geen kenmerkende veranderinge was waargeneem tussen die vinnige en stadige respons pasiënte nie. Met studie twee het die meerderheid van die selfmerkers aansienlike veranderinge oor tyd in die ongestimuleerde supernatante gewys, in vergelyking waar net die MDC en IL-4 veranderinge gedurende die observasie periode in antigeen gestimuleerde vlakke getoon het. Kenmerkende verskille is tussen die vinnige en stadige respons pasiënte in voorbehandeling vir IL-13 Ag-Nil en IL-1betaAg-Nil waargeneem. Gevolgtrekking Die studie bewys dat antigeenspesifieke response slegs beperkte potensiaal vir vroeë TB behandeling respons monitering het, maar mag die potensiaall vir onderskeidende behandeling uitkomste hê. Toekomstige ondersoeke sal dalk latere tydpunte gedurende die behandeling moet insluit aangesien dit nie in hierdie evaluasie ingesluit is nie. Die QFT-IT monsters verskyn nie as gelykwaardig met die lewendig M.tb gestimuleerde 7-dae volbloed toetse nie.

INTRODUÇÃO: A definição de fatores preditivos de lesão morfológica e funcional da tuba uterina poderia colaborar na escolha do tratamento de pacientes com gestação ectópica. O objetivo deste estudo foi verificar o comportamento do tecido trofoblástico em relação à sua penetração na parede da tuba uterina em gestações ampulares, relacionar a profundidade dessa penetração com idade gestacional, concentração de beta-hCG, tipo de imagem ultra-sonográfica e dimensão da massa ectópica à ultra-sonografia e avaliar a possibilidade de predição dessa invasão pelos parâmetros estudados. MÉTODOS: realizou-se estudo retrospectivo, entre 1° de janeiro de 2000 a 31 de março de 2004, com 105 pacientes com gestação tubária ampular submetidas à salpingectomia. As imagens ectópicas foram classificadas pelo aspecto ultra-sonográfico em anel tubário, massa complexa e embrião com atividade cardíaca e sua dimensão foi obtida pela medida do maior eixo. Histologicamente a invasão trofoblástica na parede tubária foi classificada em grau I: quando limitada à mucosa da tuba uterina; grau II: até a camada muscular; grau III: invasão de toda a espessura da tuba uterina. RESULTADOS: 29 pacientes tiveram infiltração tubária grau I, 30 pacientes infiltração grau II e 46 pacientes infiltração grau III. Os graus de invasão trofoblástica não estiveram associados à idade gestacional (p = 0,53) nem ao maior diâmetro da imagem à ultra-sonografia (p = 0,43). Os diferentes graus de invasão trofoblástica apresentaram diferença significativa da beta-hCG (p < 0,001). O grau I apresentou valores menores que os graus II e III (p < 0,05) e o grau II valores menores que o grau III (p < 0,05). Houve associação entre o grau de invasão trofoblástica e a descrição do tipo de imagem identificada à ultra-sonografia (p = 0,001). Embrião com atividade cardíaca foi mais prevalente nos casos de invasão grau III. O valor de 2 400 mUI/ml apresentou sensibilidade de 82,8%, especificidade de 85,5%, valor preditivo positivo de 68,6% e valor preditivo negativo de 92,7% (acurácia de 84,8%) para determinar invasão trofoblástica grau I. beta-hCG de 5 990 mUI/ml foi o melhor ponto de corte para predição de invasão trofoblástica grau III: sensibilidade de 82,6%, especificidade de 74,6%, valor preditivo positivo de 71,7% e valor preditivo negativo de 84,6% (acurácia de 78,1%). CONCLUSÕES: Em gestações ampulares, o tecido trofoblástico se desenvolve a partir de sua penetração na parede tubária, a profundidade da penetração do trofoblasto na tuba uterina relaciona-se às concentrações séricas de beta-hCG e ao tipo de imagem ultra-sonográfica, sendo que a concentração sérica da beta-hCG é a melhor preditora da profundidade da invasão na tuba uterina. / INTRODUCTION: The definition of predictive factors of morphologic and functional damage to the Fallopian tube may help in the choice of treatment for patients with ectopic pregnancy. The objective of the present study was to verify the presence of trophoblastic invasion into the tubal wall in ampular pregnancies, correlate the depth of penetration of trophoblastic tissue into the tubal wall with gestational age, beta-hCG concentration, type of ultrasonographic image and dimension of the ectopic mass upon ultrasound, and to evaluate the possible prediction of this invasion based on the parameters studied. METHODS: A retrospective study was conducted on 105 patients with ampular pregnancy submitted to salpingectomy between January 1, 2000 and March 31, 2004. Ectopic images were classified based on ultrasonographic findings in tubal ring, complex mass and embryonic heart activity. The dimension of the mass was determined by measuring the major axis. Histologically, trophoblastic invasion into the tubal wall was classified as grade I when limited to the tubal mucosa, grade II when reaching the muscle layer, and grade III when comprising the full thickness of the Fallopian tube. RESULTS: Twenty-nine patients had tubal infiltration grade I, 30 had grade II and 46 had grade III. The level of trophoblastic invasion was associated neither with gestational age (p = 0.53) nor with a greater diameter of the ultrasound image (p = 0.43). The different levels of trophoblastic invasion were significantly associated with beta-hCG concentration (p < 0.001), with lower concentrations being observed for grade I compared to grades II and III (p < 0.05) and for grade II compared to grade III (p < 0.05). There was an association between the level of trophoblastic invasion and the type of ultrasonographic image (p = 0.001). Embryos with heart activity were more prevalent in cases of grade III invasion. beta-hCG levels of 2 400 mIU/ml showed 82.8% sensitivity, 85.5% specificity, a positive predictive value of 68.6% and a negative predictive value of 92.7% (84.8% accuracy) for the diagnosis of grade I trophoblastic invasion. A beta-hCG titer of 5990 mIU/ml was the best cut-off for the prediction of grade III trophoblastic invasion: 82.6% sensitivity, 74.6% specificity, positive predictive value of 71.7% and negative predictive value of 84.6% (78.1% accuracy). CONCLUSIONS: trophoblastic tissue penetrate tubal wall in ampular pregnancies, the depth of penetration of trophoblastic tissue is correlated with beta-hCG concentration and type of ultrasonographic image and beta-hCG titer is the best predictor of the depth of penetration into tubal wall.

Os principais marcadores sorológicos para o diagnóstico da toxoplasmose aguda ou recente são os anticorpos IgM específicos e anticorpos IgG de baixa avidez. Entretanto em alguns pacientes, anticorpos IgM e baixa avidez de anticorpos IgG podem persistir, ultrapassando o período da fase recente aguda contribuindo para erros de interpretação diagnóstica. No presente estudo, a eficiência diagnóstica do ensaio imunoenzimático foi avaliada, com o uso de frações antigênicas ou peptídeos sintéticos originados do antígeno ESA de T.gondii, denominados de SAG-1, GRA-1 e GRA-7. Foram estudadas frações isoladas e combinadas em múltiplos peptídeos antigênicos (MAP), visando estabelecer um perfil confiável para definição sorológica de toxoplasmose recente aguda em amostra única de soro. A melhor eficiência diagnóstica do ensaio foi encontrada com o uso da combinação de peptídeos SAG- 1,GRA-1 e GRA-7, denominada MAP1. A detecção de anticorpos IgG e IgM anti- MAP1 apresentou a melhor definição entre a fase recente aguda da fase recente não aguda na toxoplasmose. Nossos resultados mostraram que IgM anti-MAP1 poderá se constituir um marcador sorológico importante no aumento da eficiência diagnóstica da toxoplasmose recente aguda / The main serological marker for the diagnosis of recent toxoplasmosis is the specific IgM antibody, along with IgG antibodies of low avidity. However, in some patients these antibodies may persist long after the acute/recent phase, contributing to misdiagnosis in suspected cases of toxoplasmosis. In the present study, the diagnostic efficiency of ELISA was evaluated, with the use of peptides derived from T. gondii ESA antigens, named SAG-1, GRA-1 and GRA-7. In the assay referred to, we studied each of these peptides individually, as well as in four different combinations, as Multiple Antigen Peptides (MAP), aiming to establish a reliable profile for the acute/recent toxoplasmosis with only one patient serum sample. The diagnostic performance of the assay using MAP1, with the combination of SAG-1, GRA-1 and GRA-7 peptides, demonstrated better discrimination of the acute/recent phase from non acute/recent phase of toxoplasmosis. Our results show that IgM antibodies to MAP1 may be useful as a serological marker, enhancing the diagnostic efficiency of the assay for acute/recent phase of toxoplasmosis

A utilização da cintilografia de perfusão miocárdica (CPM) estava limitada a pacientes que podiam realizar algum tipo de exercício, para ampliarmos a disponibilidade clínica da CPM, vários protocolos com estresses farmacológicos foram desenvolvidos. A adenosina fármaco amplamente utilizado, potente vasodilatador coronariano, apresenta uma importante interação com outras substâncias que anatagonizam seus efeitos, o dipiridamol, alimentos com cafeína e derivados de xantinas. O tempo de suspensão de cafeína da dieta para o uso exógeno da adenosina na realização da CPM ainda não está definido. Para testar essa hipótese avaliamos a influência da abstinência de cafeína em 24h, 12h e 1h, através de sua dosagem sérica, antes do estresse farmacológico com adenosina e sua possível repercussão nas imagens da CPM e o efeito vasodilatador no sistema cardiovascular. Definimos como objetivo primário: comparar a presença e a extensão dos defeitos reversíveis da CPM verificados em pacientes com abstinência de café por 24 horas (E1) com as imagens da randomização com 1 hora e 12 horas (E2) sem cafeína e como objetivos secundários: avaliar a presença e intensidade dos paraefeitos, comportamento da frequência cardíaca e da pressão arterial sistêmica. Foram submetidos ao estresse farmacológico com adenosina 194 pacientes para a realização da CPM, dos quais 43 pacientes preencheram os critérios para a randomização (defeitos perfusionais transitórios na CPM com adenosina). Excluímos seis pacientes (13,9%), três (6,9%) se recusaram a realizar a fase da randomização e os outros três (6,9%) nos quais não houve consenso entre os observadores em relação aos defeitos de perfusionais transitórios. A média de idade dos 37 pacientes analisados foi de 61,4 ± 8,3 anos, sendo 21 pacientes do sexo masculino (56,8%). Na avaliação das imagens da CPM, não houve diferença entre as imagens obtidas no grupo E1 comparadas as imagens de 1 (2,0 ± 1,5) hora ou 12 (12,6 ± 3,1) horas sem cafeína (E2). As médias de cafeína sérica encontradas foram de 0,14 ± 0,17 mg/l no grupo E1 do E2 de 1 hora e 0,13 ± 0,24 mg/l no grupo E1 do E2 de 12 horas, na randomização de 1 hora de 1,97± 0,83 mg/l e de 12 horas de 1,51 ± 1,46 mg/l (E1 vs. E2: p < 0,001). Em relação à presença dos paraefeitos, ocorreram em 31 pacientes (83,7%) e os mais freqüentes foram: dor precordial atípica, cansaço e dor em região cervical. Não foram observadas diferenças em relação às freqüências absolutas e relativas na ocorrência de paraefeitos entre os grupos. A intensidade dos paraefeitos foi verificada através de análise subjetiva comparando os sintomas em relação aos exames. Notou-se que em 22 pacientes (59,4%), caracterizaram o estudo randomizado como bem melhor, ou melhor, que o de 24 horas. A pressão arterial sistêmica e a freqüência cardíaca também não apresentaram diferenças entre os grupos. Conclusões: Os resultados permitem inferir que o uso da cafeína ingerida 2 horas antes da realização da CPM com adenosina foi eficaz e segura, pois apesar da modificação da resposta vasodilatadora máxima alcançada, traduzida pela melhor tolerância do exame realizado com menos tempo de ausência de cafeína na dieta, não modificou o resultado final da CPM / Myocardial perfusion imaging (MPI) in the past was only available to patients who were able to perform some type of exercise. Many protocols were developed with pharmacologic stress in order to enlarge the clinical availability of myocardial perfusion imaging (MPI). Adenosine is widely used and a potent coronary vasodilator, exhibits a significant interaction with other substances which antagonize its effects, such as dipyridamole, food products containing caffeine and xanthine derivatives. We found no clear consensus as to the time between caffeine ingestion and a cardiovascular adenosine stress test. To test this hypothesis we evaluated the influence of 24-hour, 12-hour and 1-hour caffeine abstinence, by assessing plasma caffeine levels before adenosine pharmacological stress, the possible repercussions on MPI images and vasodilatory effects on the cardiovascular system. The primary endpoint was to compare the presence and extent of reversible myocardial perfusion defects found in patients with 24-hour abstinence from coffee (E1), with images from patients randomized to 1-hour or 12-hour caffeine abstinence (E2). As secondary endpoints we evaluated the presence and intensity of paraeffects, as well as heart rate and systemic arterial blood pressure behavior. For this purpose, 194 patients underwent pharmacological stress with adenosine for MPI, 43 of whom fulfilled criteria for randomization (patients with transient perfusion defects detected on adenosine MPI). Six patients were excluded (13.9%), three patients (6.9%) who refused to undergo randomization, and other three (6.9%) in whom the observers could not reach consensus regarding transient perfusion defects. Mean age for the 37 patients analyzed was 61.4 ± 8.3 years, 21 patients male (56.8%). In the evaluation of myocardial perfusion images, no differences were detected between images obtained in the 24-hour E1 arm, and those from 1 (2.0 ± 1.5) hour or 12 (12.6 ± 3.1) hours randomization E2. Mean plasma caffeine level was 0.14 ± 0.17 mg/l in group E1 of the 1-hour arm E2, and 0.13 ± 0.24 mg/l in group (E1) of the 12-hour arm (E2), respectively, compared with 1.97± 0.83 mg/l, in the 1-hour, and 1.51 ± 1.46 mg/l, in the 12-hour (E2) randomized arms (E1 vs. E2: p < 0.001). In the 31 patients (83.7%) with presence of paraeffects, the most frequent were: atypical precordial pain, tiredness and pain on cervical area. There were no differences in the absolute and relative frequency in the occurrence of paraeffects between groups. However, with respect to the intensity of pareffects, a subjective analysis was undertaken, comparing symptoms of two exams. which resulted in 22 patients (59.4%) ranking the randomized study a lot better, or better than the 24-hour one. The systemic blood pressure and the heart rate behavior also did not reveal any significant differences between groups. Conclusions: The results may imply that the use of caffeine ingested 2 hours prior to the CPM with adenosine was effective and safe, for despite the change in maximal vasodilator response achieved, manifested by better tolerance of the examination with less time in the absence of caffeine in the diet, did not change the final result of the CPM

Orientador: Rita de Cássia Loiola Cordeiro / Banca: Fausto Medeires Mendes / Banca: Lourdes Aparecida Martins dos Santos Pinto / Resumo: O objetivo do presente estudo foi avaliar a efetividade da fluorescência a laser (KaVo DIAGNOdent) para a determinação da severidade de lesões de cárie oclusal em dentes permanentes in vivo e para determinação da remineralização in vitro de lesões de cárie induzidas artificialmente em esmalte de dentes bovinos. No estudo in vitro, 78 blocos de esmalte com lesões de cárie artificial (mancha branca) com valores de dureza Vickers de 38,48 l 0,85 foram submetidos ao tratamento com dentifrício fluoretado durante a ciclagem de pH, para promover a remineralização. Antes do início do experimento, após a desmineralização e após a remineralização foram realizadas análises com a fluorescência a laser e análises de microdureza superficial do esmalte. Foram encontradas diferenças significantes nas medidas de microdureza superficial (p < 0,0001) nas 3 etapas. Os valores encontrados nas leituras com a fluorescência a laser foram extremamente baixos (entre 0-5), apresentando diferença estaticamente significante somente após a remineralização. No estudo in vivo, foram selecionados 130 sítios de primeiros molares permanentes hígidos ou com presença de lesões de cárie incipientes na superfície oclusal, em crianças com idade entre 7 a 12 anos. Após profilaxia, foi realizada a inspeção visual pelo Examinador A e 23 o exame com o DIAGNOdent pelo Examinador B. Apenas os sítios com sinais sugerindo lesão de cárie foram abertos com equipamentos que permitiram preparos conservadores e a extensão da lesão (padrão-ouro) foi determinada pelo Examinador C. De acordo com os pontos de corte propostos pela KaVo (1999) e com os novos pontos de corte estabelecidos pela curva ROC, observamos uma diferença na especificidade, 0,31 e 0,85 respectivamente, considerando todas as lesões como cariadas, e um equilíbrio entre sensibilidade ...(Resumo completo, clicar acesso eletrônico abaixo). / Abstract: The purpose of this study was to evaluate the effectiveness of laser fluorescence device (DIAGNOdent) in detecting in vivo depth of occlusal caries lesions in permanent teeth and detecting in vitro remineralization of artificial caries lesions in enamel bovine teeth. On in vitro study, 78 enamel blocks with artificial caries lesions (white spot lesions) with Vickers microhardness number of 38.48 l 0.85 were submitted to dentifrice treatment during the pH-cycling, to promote remineralization. At baseline, after demineralization and after remineralization analyses were carried through with laser fluorescence device and enamel superficial microhardness analyses. Significant differences in superficial microhardness measures were found in the 3 stages (p < 0.0001). Values found in laser fluorescence readings were extremely low (between 0-5), presenting difference statistically significant only after remineralization. On in vivo study, one hundred and third sites from occlusal surfaces of sound or carious first permanent molars were selected in children, aged between 7 and 12 years. After professional cleaning, visual inspection was performed by Examiner A and laser fluorescence (DIAGNOdent) assessment by Examiner B. Only the sites presenting signals suggesting caries lesions were opened that resulted from minimal intervention and the 26 lesion extension (gold-standard) was determined by Examiner C. In accordance with cut-off points proposed by KaVo (1999) and with the new cut-off points established by ROC curves, we observe a difference in specificity, 0.31 and 0.85 respectively, considering all lesions as caries, and a balance between sensitivity (0.70) and specificity (0.87) for the new cut-off points considering dentine lesions only. Accuracy for visual inspection was ...(Complete abstract, click electronic address below). / Mestre