Treatment of cancers has been improved and new findings in research communities areconstantly found, but there are still many questions about how to treat these complex diseases.One way to treat cancer is to expose cancer cells to drugs that kill the cancer cells to a largerextent than the normal cell from the same as well as other tissue types. Different drugcompounds have diverse molecular effects on the cancer cells and to evaluate them, studies ondifferent cell lines were performed.Experiments were performed to study morphological and metabolic changes on treatedcells. Morphological changes in growing populations of MCF-7 cells and MCF-10A cellswere studied by using a phase contrast video microscopy (IncuCyte) image analysis. Changesin levels of metabolites and proteins were analyzed using two different mass spectrometricmethods. Hierarchical clustering was used to study the relationship between the collectedspectra and the most outstanding subgroup (cluster) was a set of compounds related toestrogens.There were apparent morphological differences between the two different cell lines, bothwhen untreated and after induction of apoptosis. This study shows that, when examining themetabolic patterns, there are tendencies among the substances studied to form clustersaccording their pharmacological classes. Although more studies have to be performed in thisarea it has been showed that there are possibilities to determine which pharmacological class asubstance belongs to by examining the morphological and metabolic patterns.

Background: Since the initiation of assisted reproduction techniques, several studies has been performed to improve treatment results by development of culture conditions like embryo oil and culture media used. In this study, two embryonic oils from different companies, Nidoil and Ovoil were examined.Method: In this study, 47 human embryos were used. All embryos were donated for research purposes by couples who had been treated at the clinic in Uppsala University Hospital. The embryos were divided into two groups, one group was cultured with Ovoil and the other with Nidoil.Results: There was no difference between the two oils, the embryo quality was the same in both groups.CONCLUSION: The result was expected because both oils had the same composition and purity.

Anaerobe opportunist Clostridium difficile causes the majority of hospital-acquired antibiotic-associated diarrhea. Infections can be severe because of its ability to withstand many antibiotics, to sporulate and to produce toxins (A, B and binary).     In Sundsvall Hospital C. difficile is detected with real-time PCR, which targets the sequences of toxin B, binary toxin and a regulatory gene deletion seen in the virulent ribotype 027. All positive samples are stored frozen for one month, available for further analysis or outbreak investigation. The aim of this study was to investigate if temperature and transportation time may affect the viability of C. difficile, and the PCR-result.     Frozen feces samples were cultivated, identified with MALDI-TOF and analyzed with real-time PCR after at least one month of storage. To simulate the effect of transportation time, samples were stored at 4-8°C for three and seven days before cultivation and identification. Controls were cultivated after freezing for comparison.     Ninety percent of the frozen samples contained viable C. difficile. Discrepancies between PCR-results were found for two of the oldest samples collected (six months), which turned negative. Fresh samples showed lower amount of viable C. difficile after three days (50 %) than after seven days (60 %) of storage, perhaps because of competition with other bacteria and sporulation. The frozen control group contained a higher viable amount, 75 %. The results indicate that C. difficile tolerates to be stored at low temperatures as practiced today at the laboratory. Transportation time seem to affect the outcome of cultivation, but not the PCR-result.

Leukocyte differential count in peripheral blood smear has, ever since the method was developed more than 100 years ago, been one of the most frequently used diagnostics tool in the routine hematology laboratory. The manual differential count of leukocytes using a microscope is still the standard method in most small and medium sized laboratories. Even though the method does not require any expensive instruments it comes at a high cost due to it being labor intensive and time consuming. In recent years the rapid technical advancements has led to the development of automatic or semi-automatic methods in which the leukocytes are differentiated. In this study a method comparison was made between manual leukocyte differential counts, CellaVision DM1200 Vet and Advia 2120 when analyzing 106 fresh, feline blood samples. The general agreement between results was good, especially for the most common leukocytes, such as neutrophils and lymphocytes. Results for eosinophils and monocytes had moderate agreement. The confidence intervals were generally wider when CellaVision DM1200 Vet was compared with Advia 2120, than when CellaVision DM1200 Vet was compared to the manual differential count. Despite the fact that Advia 2120 and CellaVision DM1200 Vet are both faster and often show comparable results to the manual differential count, the light microscopy will remain the gold standard for difficult samples, where there is suspicion of inflammation (band neutrophils), intracellular microorganisms, reactive lymphocytes or if the sample contains a high degree of smudge cells or artifacts.

The impact storage has on function of platelet concentrates is not completely known, although some factors have been discovered and measures have been taken to counteract them, such as adding platelet additive solution. There are several methods for analysing platelet function. In this study, the aim was to analyse change of platelet function in platelet concentrates over time and to see what effect fibrogen has on the coagulation. A technique using free oscillation rheometry (FOR), ReoRox®, was used to analyse function in platelet concentrates, both over time and after addition of fibrinogen. The platelets were analyzed at a concentration of 800 x109 Ptl/L and activated with thrombin receptor antigen peptide (TRAP). For fibrinogen efect analysis, four different concentrations were used, 10 g/L, 2,25 g/L, 1,0 g/L and 0,1 g/L. The results showed no statistically significant change in the function over time. However an increase in elasticity and decrease in the decline of elasticity could be seen. While analysing the platelets with fibrinogen it showed that up to 2,25 g/L the aggregation increased, while it decreased significantly at 10 g/L. In conclusion, the platelet concentrates retained a good clotting function from day one to day seven of storage, while the clot became stronger and fibrinolysis decreased. Fibrinogen proved important for coagulation, however a too high concentration inhibits coagulation.

Som svar på bakteriellt orsakade systemiskainfektioner och sepsis frisätts prohormonet prokalcitonin (PCT) till blodbanan.Analys av plasma-PCT (P-PCT) kan utföras med reagenset Elecsys BRAHMS PCT ianalysmodulerna cobas® e602 och e411från Roche Diagnostics.Analysprincipen är electrochemiluminiscence som bygger på immunanalys avsandwichprincip. Syftet med föreliggande studie var att verifiera metoden föranalys av P-PCT på Roche cobas® e602 och e411 med Elecsys BRAHMS PCT(ThermoFischer) samt att göra en hållbarhetsstudie av analyten ioriginalprovröret. Verifieringen gjordes genom mätning av repeterbarhet ochprecision, samt jämförelse av analysresultat från patientprover mot ett annatlaboratorium som använder samma analysmetod. Hållbarhetsstudien gjordes genomanalys av fem patientprover under olika tidsintervall 0-24 timmar efterprovtagning. Resultaten av repeterbarhetsstudien gavvariationskoefficientvärdet (CV) 3,5 % på Roche cobas® e602 medkontrollmaterial nivå 1 (åsatt värde 0,53 µg/L) och CV 1,3 % för kontrollmaterialnivå 3 (åsatt värde 24,5 µg/L). På Roche cobas e411 blev CV 2,2 % för nivå 1och 1,6 % för nivå 3. Precisionsstudien gav mellanliggande imprecisions CV-värdenmellan 1,2–1,9 % (Kontrollmaterial nivå 1 och 3). Patientjämförelserna visadeett linjärt samband (r>0,99). Bias tenderade att vara högre vidanalysresultat >20 µg/L. Hållbarhetsstudien resulterade i en obetydlig minskningav PCT koncentrationen från omedelbar analys (0,73; 57,7; 2,4; 0,94; 0,27 µg/L)och efter 24 timmar (0,70; 53,69; 2,21; 0,73; 0,24 µg/L). Repeterbarheten ochprecisionen för samtliga instrument bedömdes vara god. Patientjämförelsernavisade på ett tydligt linjärt samband med låg spridning, både vid analys iKalmar och i Linköping. Hållbarhetsstudien visade acceptabel hållbarhet avanalyten i originalprovröret. Metoden anses kunna införas på kliniskt kemiskalaboratorier vid Klinisk kemi och transfusionsmedicin, Landstinget i Kalmar län. / In response to bacterial systemic infections and sepsis, the prohormone procalcitonin (PCT) is released to the bloodstream. PCT levels in plasma can be measured using Elecsys BRAHMS PCT reagent with cobas® immunoanalyzer modules e602 and e411 from Roche Diagnostics. The test principle is electrochemiluminiscence immunoassay. The aim of this study was to verify the method for measuring plasma levels of PCT with Roche cobas® e602 and e411 using Elecsys BRAHMS PCT as well as to do a sustainability study of the analyte in the original sample tube. The verification was done by measurements of repeatability and precision, and a comparison of assay results from patient samples against another laboratory using the same method. The sustainability study was done by analyzing five patient samples during different time intervals 0-24 hours after sampling. The repeatability study gave coefficient of variations (CV %) values 3.5 % and 2.2 % with Roche cobas® e602 and 1.3 % and 1.6 % with Roche cobas® e411 using quality control level 1 (affixed value 0,53 µg/L) and level 3 (affixed value 24,5 µg/L) respectively. The precision study gave CV% values between 1.2-1.9 % (quality control level 1 and 3). The patient comparison study showed linear regression (r>0,99). Bias tended to be higher on assay results >20 µg/L. The sustainability study resulted in a slight decrease of the plasma PCT level from immediate analysis (0.73; 57.78; 2.41; 0.94; 0.27 µg/L) to analysis after 24 hours (0.70; 53.69; 2.21; 0.73; 0.24 µg/L). The repeatability and precision was considered to be good. Patient comparisons showed a clear linear relationship with little distribution between the values, both in Kalmar and in Linköping. The sustainability study showed an acceptable sustainability of the analyte in the original sample tube. This method is considered accurate and will be introduced to the laboratories of Clinical Chemistry and Transfusion Medicine at county council of Kalmar.

När leverceller drabbas av sjukdom eller cellsönderfall läcker intracellulära ämnen ut i blodet. Ämnena kan mätas på olika sätt. För att läkare ska kunna ta korrekta beslut baserade på bland annat laboratorieresultat, är det viktigt att utvärdera instrument med avseende både på reliabilitet och likvärdighet. Syftet var att jämföra Advia 1800 med Sigma Aldrichs Aspartat aminotransferas (AST) activity assay kit (AAK) avseende enzymaktivitet. I Advia 1800 analyserades 40 prover med patientserum och frystes sedan in i -20° C för att förhindra fortsatt enzymnedbrytning. Analys utfördes sedan med AST AAK med upptinade serumprov. Beräkningarna av resultaten visar att de båda metoderna har en korrelationskoefficient på 0,104 och är därmed inte jämförbara. P-värdet var <0,05 / When liver cells become subject to disease or disintegration intracellular substances will leak into the blood. These substances can be measured in different ways. For physicians to make adequate decisions based upon laboratory results, among others, it is important to evaluate instruments for both reliability and equivalence. The aim was to compare Advia 1800 with Sigma Aldrich´s Aspartate aminotransferase (AST) activity assay kit (AAK) concerning enzyme activity. In Advia 1800, 40 samples of patient serum where analyzed and then frozen at -20° C to prevent further enzyme degradation. Analysis was then performed with AST AAK on thawed samples. Results show a correlation coefficient of 0,104 and the two methods are not comparable. The p-value was <0,05

Introduction: Systemic lupus erythematosus (SLE) is an autoimmune disease that cause various inflammatory conditions in the body. The pathogenesis of this disease is yet unknown, and the diversity within the patients bring on major obstacle to clinical research for specific diagnostic markers. As a biomarker of SLE, both Interferon Regulatory Factor-5 (IRF5) and Microparticles (MP) have been suggested. Recently a study demonstrated higher concentration of IRF5+ MP in a small number of SLE patients compared to controls.  Aim: The purpose of this study was to validate and analyze IRF5+ MPs in a larger number of SLE patients and compare the results to known SLE subgroup based on IRF5 concentration.  Materials and methods: Totally 50 plasma samples from a larger cohort of SLE-patients (n=35) was analyzed together with population-based controls(n=15). Three different antibodies (in-house and commercial) were used for detection of IRF5+ MP with flow cytometry. Students t-test was used to investigate significant differences between SLE subgroup, controls and compared to the previous values. Results and Conclusion: The concentration of IRF5+ MP in SLE subgroup was significantly higher compared to controls (p<0,05). However, there were no correlations between our results and the values from the previous study, suggesting that both methods measure various forms of IRF5. These results imply that IRF5+ MP could be a possible biomarker for pathogenesis in SLE, but further studies are needed for a better understanding of IRF5, as well as of MP.

ABSTRACT The aim of this study is develop and test an inexpensive molecular tool kit to be used for diagnostic PCR for diseases such as Leber hereditary optic neuropathy (LHON) and Cystic fibrosis(CF). By developing and optimizing recombinant Taq polymerase and making a DNA size ladder from plasmids pPSU1 and pPSU2 the financial cost for the tool kit would be reduced significantly compared to the commercial components. With an inhouse method both the recombinant Taq polymerase and the pPSU1 and pPSU2 plasmids were purified from the E.coil strain DH5-α. Thereafter to analyse the components of the tool kit both conventional PCR and Real-time PCR to make sure that the tool kit would work for both types of PCRs.     The homemade Taq polymerase proved to be able to sustain in room temperature for at least 24 h and the polymerase also showed that it works with different primers such as LHON, CF and Beta-globin in both endpoint and probe base real-time PCR. The homemade size marker produced a reliable in agarose gel electrophoresis but requires optimization for continued usage for smaller PCR products.     In conclusion the homemade Taq polymerase will be used in future PCR analysis in the laboratory and the recombinant production process as well. Meanwhile the homemade size marker did not work sufficiency enough to be continuously used with gel electrophoresis in the laboratory without being further modified.

Popliteal artery entrapment syndrome has been reported to be a rare disease typically found in athletic and otherwise healthy young adults. It manifests as a temporary lower limb pain that occurs in connection with physical exercise. It is caused by an anatomic anomaly, usually anaberrant head of the gastrocnemius muscle that compresses the popliteal artery on the backside of the knee joint. The popliteal artery is the main vessel supplying blood to the lower part of the leg and the condition, if left untreated, can lead to serious complications such as thrombosis or aneurysm. If detected in time and surgically corrected patients can expect full recovery within weeks.The purpose of this study was to examine a group of healthy asymptomatic individuals (n=50)using ultrasound and a series of provocation to see if there was a possibility of entrapment. A secondary objective was to find which maneuvers during the ultrasound would provide the best results. They were subjected to ultrasound examinations at rest, during plantar flexion without resistance, against a light resistance and against substantial resistance.The results showed that most of the test subjects could temporarily constrict blood flow greatly although ultrasound imagery alone was not enough to confirm diagnosis. It seems that detected occlusion of the artery in conjunction with other diagnostic data such as AnkleBrachial Index and symptomatology can be useful both in confirming or ruling out PAESwithout the use of more expensive and invasive methods.