Neuroendocrine tumors (NET) are an unusual form of cancer that can occur in many different organs, commonly in the small intestinal. NET can be detected with the help of immunohistochemistry (IHC). IHC uses antibodies that bind to an antigen in the tissue. NET can be studied through this bond, which forms a colored product during an enzyme reaction, using a chromogenic or fluorescence-based method. Usually, NETs are analyzed with synaptophysin, chromogranin A, and CD56 markers. Novel studies have been conducted on insulin-associated protein 1(INSM1), a new emerging NET marker suggested to have higher specificity and sensitivity than the markers mentioned above.   The work aimed to investigate the optimal protocol for INSM1, test an appropriate control, and evaluate if the INSM1 antibody works according to the description in the scientific literature and is suitable to be introduced into the routine practice and used as a diagnostic marker at the department for pathology and cytology in Växjö.  The evaluation of INSM1 was performed on sectioned tissue samples with NET. NET staining was evaluated by dilution of the INSM1 antibody, different pretreatments and visualizing kits were used. The staining’s of INSM1 were assessed microscopically together with a pathologist.   The analysis showed that the optimal result was gained by using 1 mg/ml of the INSM1 antibody. The staining intensity was preferable using the OptiView compared to UltraView. The INSM1-antibodies showed better binding to the antigen when using the “standard” pretreatment and the color intensity was more intense compared to the “mild” pretreatment. The evaluation showed that OptiView and standard pretreatment, gave the optimal result for NET diagnostics using the INSM1 antibody. Based on the work herein, the antibody INSM1 could be introduced into the routine practice at the department of pathology and cytology in Växjö.

Equine Coronavirus (ECoV) is a virus that affects horses. Symptoms of ECoV are often gastrointestinal, including fever, lethargy, anorexia, and changes in fecal consistency. While the disease is usually mild, it can lead to more severe symptoms where death has occurred. Diagnosis of ECoV is typically confirmed through Polymerase Chain Reaction (PCR) testing of fecal samples. Initial studies have shown that fecal samples from horses can inhibit PCR analysis for ECoV, other PCR systems and buffers may have a better effect on inhibition in fecal samples then the current method used at Swedish Veterinary Agency (SVA) today, leading to false negative results. To avoid false negative results due to inhibition, the option of using an internal control exists. If the internal control yields a negative result, it suggests that the PCR reaction did not function as intended and a negative result for ECoV would therefore not be reliable. The purpose of this study was to gather more information about the current method's performance and possibly optimize its sensitivity. Seven different buffers were tested to see which works best for horse feces and the current method was compared with other PCR methods in order to choose the method with the highest sensitivity. The results indicated a statistically significant difference between PB and PVP buffers. Additionally the comparison between the PCR systems in the study showed a significant difference, indicating that the new system for ECoV detection has the potential to improve upon the current routine method.

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Sepsis, a life-threatening condition triggered by infection, has a long history evolving from ancient references to modern understanding. Despite advancements, its symptoms often resemble those of other illnesses until late stages, delaying recognition. The blood culture method, currently used for diagnosing sepsis, is not entirely reliable and often time-consuming, posing challenges for timely intervention in many cases. Recently, miRNAs have emerged as promising biomarkers for a variety of infectious diseases, prompting the suggestion of a new multi-marker panel for sepsis diagnosis. However, the lack of optimized methods for miRNA extraction and quantification necessitates investigation before constructing such a panel. This project aimed to examine and compare manual and robotic methods to determine which yielded superior quality, quantity, time management, and results when employing two-tailed RT-qPCR. Following experimentation, it was observed that both methods provided satisfactory purity and concentration, yet the processing time was notably shorter with the robotic method. The twotailed RT-qPCR method successfully detected DNA with copy numbers as low as 10^5 when spiked-in, although the candidate miRSeps-6 was not detectable in healthy donor plasma. Despite the unsuccessful detection of the candidate miRNA, positive controls demonstrated that processes such as manual and robotic extractions, along with two-tailed RT-qPCR, could be applied to other candidates to identify suitable matches for the kit.

Polycystic ovary syndrome (PCOS) stands out as one of the most widespread endocrine disorders affecting women, often presenting as the primary reason for infertility due to anovulation. Individuals with PCOS typically exhibit clinical and biochemical markers aligning with ultrasound findings, predisposing them to challenges such as hyperandrogenism, impaired fertility, obesity, irregular periods, excess body hair, insulin resistance, and recurrent miscarriages. The diagnosis relies on the 2003/2004 Rotterdam criteria, which identify polycystic ovary syndrome (PCOS) through the presence of two out of three key features: anovulation, clinical or biochemical signs of hyperandrogenism, and polycystic ovarian morphology. The potential underlying causes of PCOS may include genetic factors, environmental influences, or maternal imprinting. The study investigated the effect of different concentrations of dihydrotestosterone (DHT) on the oogenesis of Drosophila melanogaster. Drosophila melanogaster, widely utilized in disease research, offers valuable insights into this syndrome. With a life cycle spanning approximately 12 days, these flies undergo metamorphosis and produce the steroid hormone ecdysone, which is analogous to human testosterone. The main aim of the study is to develop a model for studying polycystic ovarian syndrome (PCOS) through the Drosophila melanogaster as an animal model. The data was collected by measuring the fecundity, Triglyceride concentration, and RT-qPCR. The results revealed no significant difference between the different concentrations of DHT and the control group. The results from fecundity, triglyceride assay, and RT-qPCR were notable but not significantly different. These findings suggest that additional experiments are necessary to confirm Drosophila melanogaster as a good model for PCOS. / <p>Det finns övrigt digitalt material (t.ex. film-, bild- eller ljudfiler) eller modeller/artefakter tillhörande examensarbetet som ska skickas till arkivet.</p><p>There are other digital material (eg film, image or audio files) or models/artifacts that belongs to the thesis and need to be archived.</p>

Diabetes, a prevalent global group of metabolic diseases, is characterized by hyperglycemia, which can lead to potential complications with prolonged exposure. Treatment goals involve lowering blood sugar levels and preventing future complications, with glycated hemoglobin, HbA1c serving as a marker for treatment control. HbA1c, forms when glucose binds to hemoglobin, aids in treatment monitoring. This study aimed to validate capillary HbA1c analysis on the Cobas Pro analyzer to comply with new in vitro diagnostic (IVD) regulations and streamline processes. Unlike the high-performance liquid chromatography method, (HPLC) to be replaced, all reagents used in the Cobas Pro are IVD-marked. Transitioning to capillary HbA1c analysis involves a change in measurement principle. The current method utilizes ion exchange chromatography with photometric detection, while the Cobas Pro employs turbidimetric inhibition immunoassay. The correlation coefficient for 31 capillary blood samples between the methods was 0.97, with most data points falling within the confidence interval. Despite obtaining a statistically significant difference from the paired ttest, the difference between the measurement values did not exceed 3 mmol/mol, which is the proposed clinical threshold. Comparison of capillary and venous blood analyzed with Cobas Pro yielded a linear correlation of 0.99, with no significant difference found by the paired ttest. A coefficient of variation (CV) under 4% was achieved for controls on the Cobas Pro. The primary advantage of transitioning the method was that it complied with IVDR regulations and enhances patient safety. Based on these results, the method is deemed safe for implementation.

Diet plays a crucial role in human health, particularly in modulating inflammation, a key factor in various chronic diseases. While several studies have explored the relationship between diet and disease, the complexity of this topic demands more extensive investigation. This study aims to examine the impact of seafood-rich diet, focusing on the fatty acid content, in modulating inflammation. The dietary intervention was similar to a Mediterranean diet, emphasizing unsaturated fats while minimizing saturated fats. Seafood and fish was prioritized as a source of polyunsaturated fatty acids. Blood biomarkers, including homocysteine, vitamin B12, folate, C-reactive protein, cholesterol, and triglycerides were analyzed before and after diet. Folate and vitamin B12 levels had increased significantly after diet, with folate increasing by 11.5 nmol/L (54% increase) and vitamin B12 by 37.25 pmol/L (9.99% increase). However, triglyceride levels experienced significant reduction by 0.32 mmol/L (29.5% decrease). Amino acid analysis indicated no significant changes. Gene expression analysis of interleukin-18, using quantitative polymerase chain reaction, was hindered by data contamination, precluding further interpretation. Tumor necrosis factor gene expression revealed no significant differences. However, the significant increase in vitamin B12 and folate levels stays an important discovery. Both molecules play essential roles in a pathway aimed at reducing homocysteine levels, which is associated with inflammation. This finding underscores the potential of seafood-rich diet in preventing inflammation. Additionally, the study lays groundwork for potential improvements in dietary guidelines as a preventative strategy against inflammatory diseases.

Stem cell niches serve as vital reservoirs, maintaining stem cells in a dormant state through specific niche components. Various biomarkers associated with stem cell niches have been identified in different tissues, including cardiac tissue, particularly at the atrioventricular junction. This study collected mouse, donor, and explanted cardiac tissue from the right side of the heart, focusing on the atrioventricular junction. Additionally, left ventricle samples from donor hearts were harvested. All samples were examined via immunohistochemistry to detect niche-related biomarkers. The study aimed to validate the atrioventricular junction as a potential niche area and to explore Cathepsin K asa possible cardiac stem cell marker, noting its slight nuclear expression. Clear expression of the stemcell marker SSEA4, the hypoxia marker Hif-1α, and the extracellular matrix marker ACAN, along with weak expression of the extracellular matrix marker CILP2, was observed, alongside Cathepsin K. Notably, explanted cardiac samples exhibited lower marker expression, except for Hif-1α, suggesting reduced oxygen levels in failing hearts. Mouse samples displayed the highest expression for each marker, mirroring the distribution seen in human hearts. Although the study does not conclusively validate the existence of a cardiac stem cell niche or the role of Cathepsin K, its findings significantly advance niche research. Further investigation is warranted to elucidate the role of the atrioventricular junction and associated biomarkers in cardiac stem cell biology.

Liver Cancer, especially hepatocellular carcinoma is a major global health challenge with increasing occurrence and mortality rates. This study aimed to explore and understand the molecular signature of the disease using bioinformatic analysis with The Cancer Genome Atlas Liver Hepatocellular Carcinoma dataset to identify differentially expressed genes and somatic mutations associated with patient survival. Transcriptome analysis revealed a significant difference in gene expression between tumor and normal tissues. Survival analysis linked specific genes to patient outcomes which suggests potential prognostic markers further proved by clinical enrichment analysis using Metascape, an online platform. Single nucleotide variant analysis uncovered frequently mutated genes, including TP53, CTNNB1, and TTN, and characterized the mutation landscape, revealing prevalent C&gt;T transitions. The integration of transcriptome and mutation data complements potential therapeutic targets and driver mutations which are important for the liver cancer development and progression. Overall this study provides valuable insights and is a framework into the molecular mechanisms of HCC and identifies potential DEGs, biomarkers (FCN2, FCN3, and COLEC10), and clinically actionable targets for improved diagnosis and therapeutic strategies.

Avian influenza, known as bird flu, is a highly contagious acute respiratory infection that infects many avian species, humans and other animals. Birds infected with Influenza A virus can shed large amounts of virus in feces, which can be detected in the lakes. Similarly, people infected with enteric viruses also shed high amounts of virus in their feces, which can end up in lake water. In this study, the presence of influenza A and seven enteric viruses (norovirus GII, astrovirus, enterovirus, adenovirus, sapovirus, hepatitis E virus, and pepper mild mottle virus) were investigated in recreational lakes to explore whether their identification could be used as an early warning of outbreaks. Lake water samples were collected from Axlemossen, Delsjön, Slottsskogen, and Torpadammen. The samples were taken monthly from November 2023 until April 2024 and analyzed for detection of viruses by concentration through ultrafiltration and quantitative polymerase chain reaction. Influenza A virus was selectively detected in the urban lakes of Slottsskogen and Torpadammen, where favorable pH, temperature, and high bird populations likely supported the virus's stability in the lakes. In contrast, enteric viruses showed a broader distribution across all lakes, likely driven by human fecal contamination from recreational activities or possibly from untreated wastewater discharges from unknown sources. The method used in this study aids in studying the presence and behavior of circulating strains in birds. Additionally, it can help understand the potential public health risks of waterborne viruses, especially those transmitted through avian or human fecal matter in the local environment.