HbA1c and glucose are the most widely used indicators for glucose control, but they havesome disadvantages. Improving the diagnosis of diabetes is always ongoing, other markersare needed as a complement when standard measurements are not sufficient. One alternativeis analysis of fructosamine, which is commercially available and inexpensive.The main aim with this study was to validate a colorimetric method for analyzingfructosamine including investigation of precision, linearity and stability. Fructosamine valueswas compared with HbA1c values with and without genetic variations in the hemoglobingene. An investigation on if serum albumin concentration affects fructosamine values wasalso performed. The colorimetric method was also compared with an enzymatic method foranalysis of frutcotsamine.Blood samples were analyzed as HbA1c on Cobas 6000 c501 and for analysis of thegenetic variants Capillarys 3 TERA was used. Plasma was collected and analyzed onMindray BS-380 as fructosamine and albumin.The methods in this study were comparable and the colorimetric method had greatprecision and linearity. The correlation between HbA1c and fructosamine was R2= 0,402.Fructosamine was not affected by genetic variations in the hemoglobin molecule and may bea useful indicator of high glucose and could replace analysis of HbA1c. Fructosamine wasnot affected by albumin. The enzymatic method was shown to correlate better with HbA1cthan the colorimetric method.In conclusion, analyzing fructosamine could be an alternative to HbA1c when patientshave genetic variants and would improve the glycemic control.

In human-assisted reproduction, embryos are cultured in an environment designed to mimic the natural environment of embryo development. To maintain the optimal culture environment, the culture media is covered with oil. Unfortunately, several studies have shown that mineral oils used in embryo cultivation contains toxic substances that have negative effects on embryo development. Before these culture oils are used in production they are washed and filtered. This procedure reduces the concentration of toxic substances to an approved level.The aim of this study was to compare three different paraffin oils effect on embryo development. The study includes two oils from Nidacon (Oil A, Oil B) and one from Vitrolife (Ovoil). To compare the effect on embryo development, time points for important embryonic development stages was noted: first division after thawing, morula, early blastocyst, blastocyst, expanded blastocyst and hatching blastocyst. In addition, blastocyst classification was done on day 5 and 6 according to Gardner and Schoolcraft´s blastocyst classification system.A total of 47 human day-2 cryopreserved embryos were divided in three groups and cultured for 4 days in EmbryoSlides overlaid with Oil A, Oil B or Ovoil respectively. The results showed no significant difference in effect on embryo development regarding morphokinetics and blastocyst classification between the three examined paraffin oils. In conclusion, the results indicated the same quality and toxicity level between the three examined paraffin oils.

The skin is the largest organ in the body. Its function is to protect the body from potential harm and to maintain homeostasis. The epidermis is the outermost layer of the skin. Stratum corneum is the outermost layer of the epidermis, composed of corneocytes and surrounding lipids. The lipids are produced by different enzymes that all play a role in the formation and function of the skin permeability barrier. Mutations in genes coding for these enzymes can lead to barrier dysfunction and could cause autosomal recessive congenital ichthyosis (ARCI). Nine genes have been identified as ARCI-causative and two of them are CYP4F22 and CERS3.   The purpose of this project was to study co-localization of CYP4F22 with CERS3 and also mutated CYP4F22 enzymes, by transfecting plasmids into HeLa and HaCaT cells and performing PLA on HEKn cells. Co-localization could indicate potential interactions and by studying these more in the future, novel treatment strategies can be developed for ARCI patients.   Transfection attempts showed a low transfection grade of wild type genes in both HeLa and HaCaT cells. Tendencies towards co-localization was seen in both cell types and some HeLa cells showed strong correlation after image analysis. Transfection of mutated genes failed, unfortunately. PLA showed co-localization in normal keratinocytes. The obtained results indicated a co-localization, but results need to be confirmed by immunoprecipitation and immunoblotting in the future.

No description available.

No description available.

För att utreda anledningen till bland annat otit och sinuit tas prov från nasofarynx, som enligt nuvarande metod utodlas på två separata plattor med blod- och hematinagar för att identifiera patogena luftvägsbakterier. Diagnostiska antibiotikadiskar används för att förenkla differentiering mellan olika bakterier. Odling på blodagar med optochin-disk är en tillförlitlig metod för identifiering av Streptococcus pneumoniae. Metoden är tidskrävande och agglutinationstest är ett alternativ för mer tidseffektiv artidentifiering. Syftet med arbetet var dels att validera en ny provsättningsmetod för nasofarynx-prov samt att utvärdera ett nytt agglutinationstest för artidentifiering av S. pneumoniae. För att validera provsättningsmetoden jämfördes den tidigare metoden med en ny där analyserna kombineras på samma platta (biplate) (N=165). För validering av nytt agglutinationstest inokulerades hästblod och bakterieisolat i blododlingsflaskor. Agglutinationstest utfördes därefter med två kommersiella kit, Dryspot, som idag används på laboratoriet, och Immulex. Fynden från nasofarynx efter odling på nya kombinerade plattor bestod av S. pneumoniae, Haemophilus influenzae och Moraxella catarrhalis. Överensstämmelsen mellan odlingsmetoderna var god för nasofarynxodlingar. Vid mätning av bakteriefri zon kring optochin-disk erhölls 18 – 31 mm för biplates respektive 16 – 24 mm för originalmetoden med odling på två separata plattor. För två prov var den bakteriefria zonen på biplates ej mätbar. Vid agglutinationstest för identifiering av S. pneumoniae erhölls agglutination inom respektive tidsgräns för samtliga referensisolat. Ett α-streptokock- och ett enterokock-isolat agglutinerade vid test med Immulex. Slutsatsen av studien är att biplates bör fungera som rutinmetod för odling av nasofarynxprov. Däremot verkar agglutinationstest med Immulex vara mindre pålitligt än Dryspot som används idag. / To investigate the cause of otitis and sinusitis, samples are taken from nasopharynx. To find airway pathogens samples are cultivated on blood agar and hematin agar, for further differentiation diagnostic antibiotic disks are used. Cultivation on blood agar with optochin disks is a reliable method to identify Streptococcus pneumoniae, although it is time-consuming. Agglutination test is a rapid method for identification of Streptococcus pneumoniae. The aim of this study was to validate a new method for cultivation of nasopharynx-samples. Moreover a new agglutination test for identification of S. pneumoniae was evaluated. To validate the new method of cultivation, the original method was compared with the new where blood and hematin agar are combined on the same plate (biplate) (N=165). To validate the new agglutination test, blood culture bottles were inoculated with blood and suspensions of relevant bacterial species. Agglutination tests were performed using Dryspot and Immulex commercial kits. Bacterial findings from nasopharynx after cultivation on the new combined plates included S. pneumoniae, Haemophilus influenzae and Moraxella catarrhalis. The correlation between the methods was good for nasopharynx-cultivations. The inhibitory zones around the optochin disk were 18 – 31 mm for biplates and 16 – 24 mm for the original method. All S. pneumoniae agglutinated within the time limits when agglutination test were performed. One α-Streptococcus and one Enterococcus agglutinated when tests were performed with Immulex, hence giving false positive results. In conclusion, the new cultivation method of nasopharynx samples proved functional.  Agglutination test with the kit presently used, Dryspot, performed better than Immulex.

Prostatacancer är idag den vanligaste cancerformen i Sverige, och drygt 10 000 män årligen blir diagnosticerade med sjukdomen. En obotlig variant med dödlig utgång är kastrationsresistent prostatacancer som behandlas primärt med Docetaxel. Tumörcellerna utvecklar dock med tiden resistens mot cytostatikan, och det föreligger därför ett stort behov av att utveckla kombinationsbehandlingar med Docetaxel som grund. Mangan har i flera studier visat sig kunna inducera apoptos hos olika celltyper, och är därför intressant som ett möjligt komplement till Docetaxelbehandling. Syftet med studien var att analysera apoptos hos prostatacancerceller från cellinjen PC3 som exponerats för Docetaxel i kombination med mangan. Cellerna odlades och behandlades med Docetaxel i kombination med mangan under 24, 48 och 72 timmar. Analys av apoptos utfördes med flödescytometer efter infärgning med Annexin V och propidiumjodid. Beroende på cellernas tillstånd färgas de in i olika kombinationer och kan därmed detekteras som tidigt respektive sent apoptotiska, eller som nekrotiska. Resultaten visade på en ökning av apoptos hos kombinationsbehandlade celler, men ingen signifikant skillnad förelåg i jämförelse med obehandlade celler. Ytterligare försök bör därför upprepas med ökad koncentration av Docetaxel i kombination med olika koncentrationer av mangan, och även med andra prostatacancer cellinjer såsom DU145 och LnCaP. Dessutom bör analyser av apoptotiska markörer genomföras, i syfte att bekräfta apoptos.

Brain ischemia events are among the first three mortalities causes in the world associated with high treatments costs and in most cases some degree of permanent functional disability, thus it is necessary with new low cost and effective treatments. In the present study the potential neuroprotective effect of magnesium sulphate was evaluated by quantification of ischemic biomarkers: fluoride resistant acid phosphatase (FRAP), inorganic phosphate (Pi), lactate dehydrogenase (LDH) and nitric oxide (NO) in rats with global ischemia. Adult rats were anaesthetized (sodium thiopental 60mg.kg-1) and magnesium sulphate (1mmol.kg-1) (9 animals) or saline solution (NaCl 0,9%) (11 animals) was infused via jugular vein. Cortical sample was extracted 30 minutes after infusion (normoxic condition), thereafter it was induced global ischemia via respiratory arrest caused by jugular administrated muscular relaxant (Flaxedil, 500μL), and 30 minutes after this state a cortical sample of contralateral hemisphere was taken (hypoxia condition). Samples were preserved with proteases inhibitors (PMFS y NaF) and homogenized. Biomarkers were quantified within 24h from the experiment by the following spectroscopic methods: Gomori (FRAP), Ammonium molybdate (Pi), Pyruvate to lactate reduction (LDH) and Griess (NO). FRAP activity was quantified as an inflammatory biomarker for the first time in the cerebral cortex. Global ischemia increased all biomarkers concentrations of the saline group. During normoxia condition magnesium sulphate reduced Pi (P=0,0002) and LDH activity (P=0,001), while during hypoxia it reduces Pi (P= 0,0002) and LDH activity (P=0,03) compared to saline values. These results strongly suggest the cortical neuroprotectiveeffects of magnesium sulphate in global ischemia induced by respiratory arrest, by reduction of cellular acidosis and energetic deficit.

Infertility is a widespread problem around the world, although the available treatment options constantly improve through research and methodological development. A common treatment option mainly for biologically caused infertility is in-vitro fertilization, where in a menstrual cycle multiple oocytes are stimulated to mature and are then fertilized in a laboratory. This often results in multiple good quality embryos which can be cryopreserved through vitrification and used in a later cycle to increase the success chance of the treatment. The purpose of this pilot study was to evaluate vitrification and warming kits from Kitazato and Irvine Scientific regarding results and procedures. Human cleavage stage embryos (n=76) were thawed and cultured to the blastocyst stage. The blastocysts were scored according to Gardner and cryopreserved with vitrification and warming kits from either Kitazato (n=20) or Irvine Scientific (n=20). The warmed blastocysts were controlled after 2 and 4 hours for re-expansion and freeze injuries. The data was analysed with Fisher’s exact test and considered statistically significant if two-tailed p-value <0.05. The results showed no significant difference between the kits after 2 respectively 4 hours regarding re-expansion (p=0.432; p=0.492) or freeze injury (p=1.000; p=0.476). A significant difference was observed between group AB (with higher Gardner-score) and group C (with lower Gardner-score) in the degree of freeze injury (p=0.048; p=0.034), regardless of vitrification kit used. The details of the procedures differed somewhat between the kits, both having pros and cons, although overall procedures were equivalent. Further evaluation is needed before a change in method can be conducted.

Paired box (PAX) gener kodar för en familj av nio stycken PAX transkriptionsfaktorer. PAX transkriptionsfaktorer är uppbyggda av en N-terminal DNA-bindande domän som utgörs av 128 aminosyror, en oktapeptid, och en C-terminal DNA-bindande domän. Studier har visat att PAX8 har en betydande roll vid utvecklingen av olika typer av tumörer. MRQ-50 är en antikropp som används inom Region Skåne för att påvisa PAX8 uttryck vid histopatologisk diagnostik. MRQ-50 riktas mot N-terminalen av PAX8 som är homolog med N-terminalen av PAX5. Detta kan leda till en korsreaktion i B-lymfocyter samt neoplasier, vilket inte är optimalt för diagnostik. Av den anledningen fortsätter forskning för att hitta en ny klon som kan ersätta MRQ-50. EP331 är en antikropp som är anpassad för detektion av det nukleära antigenet PAX8 och korresponderar till C-terminalen av human PAX8 protein. Syftet med arbetet är att optimera ett immunhistokemiskt färgningsprotokoll för detektion av PAX8 med hjälp av anti-PAX8 antikroppen EP331 och undersöka om godkända resultat kan erhållas. Detta gjordes för att ta reda på om det går att optimera ett protokoll med EP331, och om det erhålls lämpligare resultat än MRQ-50. Optimeringen gjordes på kontrollvävnader njure, tuba/äggledare och appendix och testades slutligen på patientprov med tumörvävnad. För protokolloptimering testades heat induced epitope retrieval, inkuberings tid av EP331, amplifiering, Ultra Wash i olika testomgångar. Resultatet visade att kärnor i epitelceller i proximala-, distala-njurtubuli och parietala epitelceller som klär Bowmans kapsel infärgades måttligt till starkt. De sekretoriska epitelen i tuba infärgades med stark intensitet och cilierade epitel måttligt. Infärgningen på patientprover visar godkänd specificitet, med en generell måttlig till stark intensitet. Med fler forskningsstudier av EP331 är det möjligt att primärantikroppen kan ersätta MRQ-50.