Akut hjärtinfarkt är globalt sett en vanlig dödsorsak. Hjärtinfarkter behöver inte ge tydliga symptom och dödsorsaken kan därmed vara okänd fram till en klinisk obduktion och att histopatolgiska studier av hjärtmuskelvävnad genomförs. Fosforwolframsyra-hematoxylin (PTAH) är en färgningsmetod som kan användas vid visualisering av hjärtinfarkt genom att färga myokardceller, fibrin, kontraktionsband och cellkärnor blå medan kollagen färgas rosa till rödbrunt. Syftet med examensarbetet var att optimera PTAH-färgningsprotokoll för att underlätta diagnosticering av hjärtinfarkt. Tre olika färgningsprotokoll, två med PTAH med Mallory Bleach (#1 och #2) och ett med PTAH med refixering i Bouins lösning (#3), jämfördes vid inkubering över natt i rumstemperatur samt 3-4 h i värmeskåp vid 56 ºC. Vävnader som färgades in var från paraffininbäddade klossar vilka tillhörde utsvarade fall med konstaterad hjärtinfarkt. Dessa snittades och värmdes fast på oladdade samt laddade objektglas. Efter färgning graderades infärgning efter kategorierna vävnadsdifferentiering och cellkomponenter med en poängskala från 0, ej bedömbar, till 3, optimal. Det protokoll med högst poäng optimerades för att hitta bästa inkuberingstid i PTAH-lösning, därefter implementerades protokollet genom att bekräfta korrekt infärgning vid upprepade infärgningar av hjärtmuskelvävnad. Protokoll #3 visade på högst poäng efter infärgning vid jämförelsen med protokoll #1 och #2 och under optimering framgick inkubering under 4 h i värmeskåp vid 56 ºC som mest gynnsam, då denna inkuberingstid även visade på mindre känslighet vid dehydrering. Slutsatsen blev att PTAH-färgning efter protokoll #3 med refixering i Bouins lösning med 4 h i värmeskåp gav bäst resultat, men färgnyansen varierar med tid mellan dödsfallet och obduktionen. / Acute myocardial infarction (MI) is a common cause of death globally. MI without symptoms or with atypical symptoms is usually first detected at clinical autopsy with histological analyses of the myocardium. Phosphtungstic acid haematoxylin (PTAH) is a method to visualize MI by staining myocytes, fibrin, contraction bands and nuclei blue while collagen stains reddish brown. The aim of this degree project is to optimize PTAH staining protocol for MI diagnosis. Three different staining protocols, two protocols with PTAH staining and Mallory Bleach solution and one protocol with PTAH staining with refixation in Bouin’s solution (#1-3), where compared along with incubation overnight in room temperature and at different hours in heat (56 ºC). Paraffin embedded tissues from myocardium from different autopsy cases diagnosed with MI were cut before mounting and heating on non-charged as well as charged slides. After staining, results were evaluated using scores 0-3 for two parameters; differences in tissues and cell components. Highest evaluated protocol where optimized to find the ultimate incubation in PTAH staining solution. For implementations, additional myocardial tissues from other cases were stained to confirm repeated results. Protocol #3 was evaluated higher in comparison with #1-2 and optimized to 4 h incubation in PTAH-solution in heating with staining results less sensitive to dehydration. In conclusion, staining protocol #3 with 4 h heating was optimal, however, vulnerable to prolonged morgue storage time losing the insensitivity of colour.

Heart failure with preserved ejection fraction has its main pathophysiology in the diastolic dysfunction which is characterized by increased filling pressures. This can be discovered during a routinely performed echocardiographic examination. In 2016 the American Society of Echocardiography published a recommendation to simplify the assessment of diastolic dysfunction. Whether or not these recommendations have simplified the gradings is questionable. Left atrial strain has been shown to decrease in the absent of diastolic dysfunction and 3D echocardiographic volume measurements of the left atrium has been showing more accurate volumes of the heart compared to 2D-measurements. A quantitative study was therefore made to compare the results from the recommended algorithm with the two methods - left atrial strain and 3D-measurements of the left atrium volume. A total of 138 women and men were included with the data collected both prospective and retrospective. The aim with this quantitative study was to examine if LA-strain and 3D volume measurements of the left atrium can be useful parameters to use in detecting diastolic dysfunction in future recommendations. The result showed significant differences (p<0.05) in LA-strain between normal diastolic function and grade 2 and -3. Also, between grade 1 and grade 2- and 3. Between undetermined diastolic function there was a statistical difference with grade 2- and 3. LA-strain showed a significant negative correlation between E/é and 2D. LA-strain and 3D-measurements of the left atrium can be useful tools in the next recommendations of grading diastolic dysfunction.

Neuroborrelios (NB) kan förekomma när Borrelia garinii, som är en del av artkomplexet Borrelia burgdorferi sensu lato (s.l), infekterar nervsystemet. När immuncellerna utsätts för borreliaantigen bildas antikroppar i plasma och intratekalt. NB ger även upphov till mononukleär pleocytos i cerebrospinalvätska (CSV). Neurologiska symptom, pleocytos och antikroppsindex (AI), som indikerar intratekal syntes av borreliaantikroppar, vägleder nuvarande NB-diagnostik på Klinisk Mikrobiologi i Kalmar. Diagnostiken försvåras vid tidig NB när AI är negativ och pleocytos positiv. För att komplettera tidig NB-diagnostik kan kemokinet C-X-C motif ligand 13 (CXCL13) mätas i CSV. I nuläget utförs CXCL13 enzymkopplad immunadsorberande analys (ELISA) på ett externt laboratorium. Mätning av CXCL13 kan även utföras med en snabb semi-kvantitativ ReaScan+ CXCL13 metod som bygger på lateral flödesimmunanalys (LFIA). Syftet var att verifiera ReaScan+ CXCL13, kontrollera linjäritet, CXCL13 stabilitet och bestämma diagnostisk specificitet och sensitivitet. CSV-proverna indelades i fyra grupper baserat på pleocytos och AI samt tre kategorier utifrån kriterier till NB-diagnos (n=31). Metodjämförelsen genomfördes mellan ReaScan+ LFIA och CXCL13 ELISA (n=25). Linjäritetsanalys utfördes på prov med hög CXCL13-koncentration i CSV och CXCL13-stabiliteten kontrollerades på två patientprover vid frys- och kylförvaring. Beräkning av sensitivitet och specificitet baserades på definitiv (n=20) och ej NB (n=5). Metodjämförelsen visade en fullständig överensstämmelse och Spearman´s korrelationskoefficienten var 0,949. Determinationskoefficienten för linjäritetsanalysen var 0,963. Diagnostisk sensitivitet var 80% och specificitet 100%. Hög prestanda, acceptabla stabilitets- och linjäritetsresultat samt ReaScan+ potentiella förmågan att särskilja mellan definitiv och ej NB, indikerar att metoden kan införas på Klinisk Mikrobiologi i Kalmar och komplettera nuvarande diagnostik av tidig NB. / Lyme neuroborreliosis (LNB) occurs when Borrelia garinii, part of the Borrelia burgdorferi sensu lato (s.l) species complex, infects the nervous system. Upon exposure, immune cells produce antibodies that can be measured in serum or cerebrospinal fluid (CSF). LNB also cause mononuclear pleocytosis in CSF. Neurological symptoms, pleocytosis and antibody index (AI), which indicate intrathecal synthesis of Borrelia antibodies, guide current diagnosis of LNB at the Clinical Microbiology in Kalmar. The diagnosis is complex in early LNB, especially when AI is negative and pleocytosis is positive. Expression of chemokine C-X-C motif ligand 13 (CXCL13) in CSF can be used as an additional marker to early LNB. Analysis is currently performed at an external laboratory using enzyme-linked immunosorbent assay (ELISA). The aim of this project was to evaluate the performance of the rapid semi-quantitative lateral flow immunoassay (LFIA) ReaScan+ CXCL13 test using 31 patient samples divided into four groups based on pleocytosis and AI. Linearity assay was performed on samples with high CXCL13 concentrations and stability was tested on two CSF samples after freezing and refrigeration. Estimation of sensitivity and specificity was based on definite (n=20) and non-LNB (n=5). Method comparison showed a complete agreement and Spearman's correlation coefficient was 0,949. The R2 for linearity was 0,963. Sensitivity was 80% and specificity 100%. Strong agreement, acceptable stability and linearity results as well as ReaScan+ potential ability to distinguish between definite and non-NB, indicate that the method can be introduced at Clinical Microbiology in Kalmar and used as a supplement in early LNB diagnostics.

In this study, the efficacy of MarrowMAX, a new IVD-certified culture medium for bone marrow samples, was evaluated and compared to the in-house modified culture medium RPMI-1640, which is routinely used at the Academic Hospital in Uppsala for chromosomal analysis. The hypothesis was that the new culture medium MarrowMAX would lead to an increased number of metaphases, resulting in a higher mitotic index, better spreading, and improved metaphase morphology. Twenty patient samples were cultured in parallel using both media and subjected to identical methods for sample preparation and staining for chromosomal analysis. The results demonstrated significant differences between MarrowMAX and RPMI-1640 for one-day, two-day, and three-day cultures. However, no significant differences were observed for directly harvested cultures and overnight colcemid cultures. These findings suggest that MarrowMAX is more effective than RPMI-1640, resulting in a higher frequency of metaphases, which is crucial for detailed chromosomal analysis. This was further confirmed by flow cytometry analysis, showing a higher number of proliferated cells in the MarrowMAX culture compared to the RPMI-1640 culture, in one representative sample. MarrowMAX's enhanced capacity to promote cell growth and structure in a laboratory setting indicates its potential as a promising alternative to RPMI-1640 for future diagnostic applications, potentially leading to faster diagnoses and improved treatment efficacy for patients with hematological conditions.

Efter upptäckten av penicillinet observerades en ökad resistens hos S. aureus vilket ledde till att alla stammar ansågs vara resistenta. Dock har många stammar enligt senare studier visat sig vara känsliga vilket har öppnat upp möjlighet att åter använda benzylpenicillin som ett behandlingsalternativ eftersom det kan ges i lägre koncentration jämfört med andra alternativ. Metoden för att upptäcka penicillinresistens hos S. aureus enligt EUCAST har bedömts svår att tillämpa på grund av subjektivitet vid bedömning av zonkanten. Studiens syfte var att utvärdera metoden för senare implementering på Klinisk Mikrobiologi Unilabs i Skövde. Metodens användbarhet utvärderades genom resistensbestämning av en referensstam samt tio stammar från ett referenslaboratorium för EUCAST. Bedömningen av de tio stammarna från referenslaboratoriet stämde vid jämförelse överens med deras resultat förutom för en stam som bedömdes som resistent i stället för känslig. Repeterbarheten var hög då ingen skillnad (p=0,156) uppvisades för referensstammen.  Resultatet tyder på att det kan vara svårt att göra en korrekt tolkning av zonkantens utseende då det är en subjektiv bedömning. För att uppnå en högre säkerhet kan det vara fördelaktigt att mer än en person gör en bedömning. Slutsatsen var att metoden kan implementeras på laboratoriet. / Following the discovery of penicillin, resistance rates increased until all strains of S. aureus were considered resistant. Recent studies have shown that many strains are susceptible, making it possible to again use penicillin as an alternative for treatment since it can be used in lower concentrations compared to alternatives. The EUCAST method for discovering penicillin resistance in S. aureus has been judged difficult due to the subjectiveness of assessing the zone edge. The aim of this study was to evaluate the method to implement it in the Section for Clinical Microbiology Unilabs Skövde. The useability of the method was evaluated by resistance testing of a reference strain and ten strains from a EUCAST reference laboratory. The assessment of the strains from the reference laboratory equaled their results except for one strain that was assessed resistant rather than sensitive. The repeatability was high since no difference (p=0,156) was found for the reference strain. The results show how difficult it is to correctly assess the zone edge due to its subjectiveness. It could be beneficial with more than one person assessing the zone edge to achieve a higher security. In conclusion the method is possible to implement in the laboratory.

Infertility is a global challenge, often remedied with In vitro fertilization (IVF) and Intra cytoplasmic sperm injection (ICSI). Sperm quality is crucial, prompting ICSI when compromised. Routine sperm preparation via gradient centrifugation raises concerns about sperm stress and DNA fragmentation. Zymot, a new device that utilizes microfluidic technology, emerges as a promising alternative. It minimizes sperm stress and DNA damage, potentially enhancing fertilization rates and embryo quality. The aim of this study was to compare the outcome of ICSI samples treated with Zymot against gradient centrifuged samples. Focus was on fertilization rates, embryo quality, and pregnancy outcomes.  The results of 104 Zymot treated samples from men with compromised sperm quality were compared with 144 gradient centrifugations retrospective. Results revealed a significant difference between methods in the number of pronulear (PN), specifically in the Good Quality Embryo (GQE) where 62% with Zymot were 2PN compared to 59% with gradient (p=0.017). No significant difference in pregnancy rates or embryo utilization rate were observed. A tendency towards higher proportion (54.6%) of Zymot-treated embryos were cryopreserved compared to gradient (49.4%, p=0,27). In conclusion, a significant difference between methods in the GQE proportion of 2PN embryos favored Zymot. Closer examination revealed a higher proportion of embryos cryopreserved with Zymot, suggesting a potential for increased treatment success in future cycles. Zymot, requiring less time, yielded equivalent results to gradient centrifugation, with higher GQE proportions and more embryos cryopreserved. This merits consideration as a high-quality alternative to sperm preparation for ICSI in cases of poor sperm quality.

År 2020 stod lungcancer för högst andel cancer-relaterade dödsfall. Cellblockstekniken används inom diagnostik av vätskebaserad cytologi för att morfologiskt undersöka cellernas utseende och tumörcellers ursprung. I dagsläget upplever personal på Avdelningen för Klinisk patologi och cytologi Jönköping problematik vid klotformering av cellerna, vilket kan resultera i cellförlust. Syftet var att testa en ny metod för preparering av cellblock för lungcytologi samt utvärdera immunohistokemisk färgning med metoden. Studien behandlade 20 prover av pleuravätska och 20 lungcytologiprover som bearbetades till cellblock. Proteinglycerol och formaldehyd ersatte nuvarande agglutineringslösning bestående av koncentrerad formaldehyd och ättiksyra. Cellmängd beräknades på 39 snitt av två cytodiagnostiker, och graderades i tre nivåer från sparsam till riklig (1–3). Åtta cellblock selekterades för immunohistokemisk färgning. Det totala medelvärdet för cellularitet på snitten var måttlig (2,04). För pleuraproverna var medelvärdet 1,775 och för lungcytologiprover 2,315. Inga celler hittades på n=4 prover. Samtliga (n=8) immunohistokemiska färgningar med tillhörande kontroller godkändes av metodspecialist. Metoden ger snitt med tillräcklig cellularitet som fungerar väl med immunohistokemisk färgning. Metoden kan optimeras i relation till proteinförhållande och provtagning för att vid implementering bidra till en effektiviserad och individualiserad diagnostik. / In 2020, lung cancer accounted for the highest proportion of cancer-related mortalities. The cell-block technique is used in liquid-based cytology to morphologically examine the appearance of cells and the origin of tumour-cells.  Staff at the Department of Clinical pathology and cytology in Jönköping are experiencing problems with clot-formation of cells, which can result in cell-loss. The aim was to test a new method for preparing cell-blocks for lung cytology, and evaluate immunohistochemical staining with the method. The study treated 20 pleural fluids and 20 fine needle aspirations that were processed into cell-blocks. Protein-glycerol and formaldehyde replaced the current agglutination solution consisting of concentrated formaldehyde and acetic acid. The cell-count on 39 sections were graded by two Cytotechnologists in three levels from sparse to abundant (1-3). Eight cell-blocks were selected for immunohistochemical staining. The mean cellularity was moderate (2,04). For the pleural and lung cytology samples the mean values were 1,775 and 2.315. No cells were found on n=4 sections. Every (n=8) immunohistochemical staining with associated controls was approved by method specialist. The method provides sufficient cellularity and works with immunohistochemical staining. It can be optimized in relation to protein-ratio and sampling to contribute to streamlined and individualized diagnosis if implemented.

Blod består av olika komponenter; erytrocyter, trombocyter, leukocyter och plasma.Plasma är en proteinrik vätska som bland annat innehåller koagulationsfaktorer. Vidberedning av donerat helblod kan dessa blodkomponenter separeras från varandra,för att sedan bearbetas vidare och-, eller användas för komponenten relevantaändamål. Plasma kan användas till transfusion vid bland annatkoagulationsrubbningar, alternativt användas som råvara förläkemedelsframställning. För att säkerställa kvaliteten på framställd plasma utförskomponentkontroller, som säkerställer att framställningsprocesser leder till önskvärtresultat. Plasma kontrolleras för innehåll av erytrocyter, trombocyter och leukocyter.Olika analysmetoder kan användas för detta, där BD Plasma Count Kit i dagslägetanvänds för att bestämma leukocytpartikelkoncentration (LPK),trombocytpartikelkoncentration (TPK) och erytrocytpartikelkoncentration (EPK).Sysmex XN Blood Bank mode (BB-mode) är en mjukvara som möjliggör analys avkomponentkontroller, och är i jämförelse en mindre resurskrävande (personal, tid,ekonomi) metod. Syftet med arbetet var därför att utvärdera denna metod;undersöka preanalytiska faktorer, verifiera provstabilitet och jämföra mednuvarande metod (BD Plasma Count Kit). Totalt 50 plasmakontroller analyseradesmed båda metoderna, till två av dessa kontroller tillsattes leukocyter.Reproducerbarhet för BB-mode mättes vid normal nivå och högre nivå, provernaanalyserades 10 gånger på varandra följda mätningar. Regressionsanalys förnuvarande metod och BB-mode utfördes för LPK, TPK och EPK. Svag korrelationförelåg för LPK (R2=0,8232), god korrelation för TPK (R2=0,9648) och dåligkorrelation för EPK (R2=0,5211). Analys av komponentkontroller (LPK, TPK ochEPK) med BB-mode gav inte samma resultat som BD Plasma Count Kit(signifikansnivå 0,05). BB-mode visar potential till förenklad analys avkomponentkontroller för plasma, men fler studier krävs. / Blood can be divided into different components: erythrocytes, thrombocytes,leucocytes, and plasma. Plasma is a fluid with proteins suspended in it. Some ofthese proteins Is coagulation factors, which makes plasma a viable treatment optionfor coagulation disorders. It could also be used for pharmaceutical production,where said coagulation factors is the main target. Quality controls (QC) of theproduced plasma ensure that the processes lead to the desired results. Residual cellsin plasma are used, and concentrations of erythrocytes, thrombocytes and leucocytesis measured in plasma. BD Plasma Count Kit is the method used today, but a newaddition to the haematology analysers Sysmex XN-series called Sysmex XN BloodBank mode (BB-mode) aims to analyse QC for blood products. The method wouldbe less resource consuming (time, personnel, economy) than the method used today.Therefore, the aim of this study is to evaluate analysis of quality controls for plasmaon BB-mode. This includes a comparison with BD Plasma Count Kit, as well asinvestigation of pre-analytical factors. A total of 50 samples were comparedbetween the methods, two of which were leucocyte spiked. Regression analysisshowed weak correlation between the methods for leucocyte concentration(R2=0,8232), good correlation for thrombocyte concentration (R2=0,9648), and poorcorrelation for erythrocyte concentration (R2=0,5211). However, the methods didnot show the same results for neither of the parameters (significance 0,05). BBmodeshow potential to use for QC on plasma, but further testing is needed.

Background: Ice hockey is a fast-paced sport with a high possibility of acquiring dental damage and concussions which can be prevented by mouthguards (MG). Despite this, many athletes choose not to use MG due to drawbacks such as discomfort and restricted airways. Restricted airways provided by MG might be reduced by external nasal dilator strips (ENDS). ENDS could contribute to increased aerobic capacity, blood oxygen saturation and self-reported compliance of ENDS by elevating airflow through the nose. Therefore, ENDS could contribute to enhanced performance and recovery rate while reducing the discomfort of MG that could lead to an increased usage of MG among healthy young adult ice hockey players. Aim: The aim was to study aerobic capacity, and oxygen saturation using mouthguard with and without external nasal dilator strips during submaximal bicycle tests. A second aim was to study the compliance of external nasal dilator strips when using mouthguard in healthy young adult ice hockey players. Method: In this experimental study using a cross-over design 15 young adult ice hockey players were contacted to attend on two separate test occasions, with and without ENDS. Aerobic capacity was measured with a Ekblom-Bak submaximal bicycle test (VO2max) and a pulse oximeter was used to examine oxygen saturation (%SpO2). Post-testing, the participants stated self-reported compliance using ENDS based on the numeric rating scale (0-10, positive-negative attitude of usage). Data are presented as mean and standard deviation. Differences between groups were analyzed with the Wilcoxon- signed rank test based on median (Q1-Q3). Result: Nine participants, aged 18-19 completed both test occasions. This study found that ENDS provided no significant difference in estimated VO2max mean (±SD), with ENDS 63.3 (±5.5) and without ENDS 64.2 (±5.0), nor %SpO2 during standard load. A significant increase showed in %SpO2 for higher individual load by 3%, with ENDS 97.1 (±1.3) and without ENDS 94.1 (±4.5). The participants experienced the test as more strenuous with ENDS than without ENDS applied, without significant difference. Despite this, the participants had a positive attitude towards the usage of ENDS. Conclusion This study regarding healthy young adult ice hockey players showed that ENDS provided no significant difference in aerobic capacity. Oxygen saturation however resulted in a positive significant difference at the higher individual load. The majority of the participants in this study propose a positive compliance with ENDS. The results are interpreted with caution due to the small sample.

Autoimmunitet är ett tillstånd där kroppens immunsystem felaktigt attackerar och skadar sina egna vävnader och celler. HLA-B27 är en genvariation som kan kopplas till autoimmun sjukdom som ankyloserande spondylit med en prevalens på 2-4% i världens befolkning. Denna studie syftade till att utvärdera och jämföra två kit för HLA-B27 alleler mot den nuvarande metoden på Länssjukhuset Ryhov i Region Jönköpings län. De metodprinciper som användes var realtids-PCR samt LAMP.  Totalt analyserades 37 avidentifierade blodprover med vardera av kiten samt med nuvarande metod. Resultatet visade en överensstämmelse med avseende på förväntade positiva och negativa resultat för HLA-B27 för de två kiten jämfört med nuvarande metoden. De tre metoderna/kiten detekterar de vanligaste HLA-B27 allelerna. Utifrån studiens resultat visade sig båda kiten vara effektiva, lättanvända samt ha stabila reagenser. Dessutom uppnådde båda kiten de IVD-krav som ställs inom EU. / Autoimmunity is a condition where the body's immune system mistakenly attacks and damages its own tissues and cells. HLA-B27 is a genetic variation that can be linked to autoimmune diseases such as ankylosing spondylitis, with a prevalence of 2-4% in the world’s population. This study aimed to evaluate and compare two kits for HLA-B27 alleles against the current method at Ryhov County Hospital in Region Jönköping County. The methodological principles used were real-time PCR and LAMP. A total of 37 anonymized blood samples were analyzed using each of the kits and the routine method. The results showed concordance with the expected positive and negative results for HLA-B27 between the two kits compared to the current method. The three methods/kits detect the most common HLA-B27 alleles.  Based on the study’s results, both kits proved to be effective, user friendly, and have stable reagents. Additionally, both kits met the IVD requirements set within the EU.