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| 2. |
- Hulteberg, Christian, et al.
(author)
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Frågeställningar i examensarbeten
- 2014
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In: [Host publication title missing].
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Conference paper (other academic/artistic)abstract
- För att klara examinationsmålen i civilingenjörsexamen (SFS 1993:100 bilaga 2) ska varje student visa förmåga att med helhetssyn kritiskt, självständigt och kreativt identifiera, formulera och hantera komplexa frågeställningar. För att öka förståelsen för hur forskningsfrågan identifieras, formuleras och hanteras vid LTH har ett tjugotal examensarbetsrapporter från olika institutioner granskats. Granskningen har utgått från rapporternas inledande och avslutande kapitel för att se om forskningsfrågan tydligt formuleras i inledningen och om den besvaras i slutsatserna. Utifrån det material vi har studerat och publicerad litteratur kan vi konstatera att det är ett allmänt problem att studenter på masters-nivå överlag har liten vana vid att hantera komplexa frågeställningar. Det tycks finnas bristande kunskaper om vad vetenskaplighet/vetenskaplig metodik/vetenskaplig tradition innebär för vad som ska presenteras och vi kan konstatera att det både på LTH och i stort finns ett behov för åtgärder för att på ett mer effektiv sätt träna studenter i detta. Att litteraturen inom detta område är begränsad visar på både behovet men kanske också svårigheter att identifiera precis hur detta kan göras. Vi identifierade detta som ett viktigt utvecklingsområde för handledare av examensarbeten inom civilingenjörsutbildningarna på LTH, som dock också kan komma att behöva stöd av nya strukturer och systematiska åtgärder för att nå gemensamma mål för hela fakulteten.
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| 3. |
- Nordenfelt, Pontus, et al.
(author)
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Antibody orientation at bacterial surfaces is related to invasive infection.
- 2012
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In: Journal of Experimental Medicine. - : Rockefeller University Press. - 1540-9538 .- 0022-1007. ; 209:13, s. 2367-2381
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Journal article (peer-reviewed)abstract
- Several of the most significant bacterial pathogens in humans, including Streptococcus pyogenes, express surface proteins that bind IgG antibodies via their fragment crystallizable (Fc) region, and the dogma is that this protects the bacteria against phagocytic killing in blood. However, analysis of samples from a patient with invasive S. pyogenes infection revealed dramatic differences in the presence and orientation of IgG antibodies at the surface of bacteria from different sites. In the throat, IgG was mostly bound to the bacterial surface via Fc, whereas in the blood IgG was mostly bound via fragment antigen-binding (Fab). In infected and necrotic tissue, the Fc-binding proteins were removed from the bacterial surface. Further investigation showed that efficient bacterial IgGFc-binding occurs only in IgG-poor environments, such as saliva. As a consequence, the bacteria are protected against phagocytic killing, whereas in blood plasma where the concentration of IgG is high, the antibodies preferentially bind via Fab, facilitating opsonization and bacterial killing. IgG-poor environments represent the natural habitat for IgGFc-binding bacteria, and IgGFc-binding proteins may have evolved to execute their function in such environments. The lack of protection in plasma also helps to explain why cases of severe invasive infections with IgGFc-binding bacteria are so rare compared with superficial and uncomplicated infections.
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| 4. |
- Olsson, Niclas, et al.
(author)
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Grading breast cancer tissues using molecular portraits.
- 2013
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In: Molecular & Cellular Proteomics. - 1535-9484. ; 12:12, s. 3612-3623
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Journal article (peer-reviewed)abstract
- Tumor progression and prognosis of breast cancer patients is difficult to assess using current clinical and laboratory parameters, where a pathological grading is indicative of tumor aggressiveness. This grading is based on assessment of nuclear grade, tubule formation, and mitotic rate. We report here the first protein signatures associated with histological grades of breast cancer, using a novel affinity proteomics approach. We profiled 52 breast cancer tissue samples, by combining nine antibodies and label-free LC-MS/MS, which generated detailed quantified proteomic maps representing 1,388 proteins. The results showed that we could define in-depth molecular portraits of histologically graded breast cancer tumors. Consequently, a 49-plex candidate tissue protein signature was defined that discriminated between histological grade 1, 2, and 3 of breast cancer tumors with high accuracy. Highly biologically relevant proteins were identified, and the differentially expressed proteins indicated further support for the current hypothesis regarding remodeling of tumor microenvironment during tumor progression. The protein signature was corroborated using meta-analysis of transcriptional profiling data from an independent patient cohort. In addition, the potential for using the markers to estimate the risk of distant metastasis free survival was also indicated. Taken together, these molecular portraits could pave the way for improved classification and prognostication of breast cancer.
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| 5. |
- Scholz, Birger, et al.
(author)
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Impact of Temperature Dependent Sampling Procedures in Proteomics and Peptidomics : A Characterization of the Liver and Pancreas Post Mortem Degradome
- 2011
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In: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 10:3, s. M900229MCP200-
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Journal article (peer-reviewed)abstract
- Little is known about the nature of post mortem degradation of proteins and peptides on a global level, the so-called degradome. This is especially true for nonneural tissues. Degradome properties in relation to sampling procedures on different tissues are of great importance for the studies of, for instance, post translational modifications and/or the establishment of clinical biobanks. Here, snap freezing of fresh(< 2 min post mortem time) mouse liver and pancreas tissue is compared with rapid heat stabilization with regard to effects on the proteome (using two-dimensional differential in-gel electrophoresis) and peptidome (using label free liquid chromatography). We report several proteins and peptides that exhibit heightened degradation sensitivity, for instance superoxide dismutase in liver, and peptidyl-prolyl cis-trans isomerase and insulin C-peptides in pancreas. Tissue sampling based on snap freezing produces a greater amount of degradation products and lower levels of endogenous peptides than rapid heat stabilization. We also demonstrate that solely snap freezing related degradation can be attenuated by subsequent heat stabilization. We conclude that tissue sampling involving a rapid heat stabilization step is preferable to freezing with regard to proteomic and peptidomic sample quality.
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| 6. |
- Säll, Anna, et al.
(author)
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AFFIRM – a multiplexed immunoaffinity platform that combines recombinant antibody fragments and LC-SRM analysis
- 2014
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In: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 13:12, s. 5837-5847
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Journal article (peer-reviewed)abstract
- Targeted measurements of low abundance proteins in complex mixtures are in high demanded in many areas, not the least in clinical applications measuring biomarkers. We here present the novel platform AFFIRM (AFFInity sRM) that utilizes the power of antibody fragments (scFv) to efficiently enrich for target proteins from a complex background and the exquisite specificity of an SRM-MS based detection. To demonstrate the ability of AFFIRM, three target proteins of interest were measured in a serum background in single- and multiplexed experiments in a concentration range of 5-1000 ng/ml. Linear responses were demonstrated down to low ng/ml concentrations with high reproducibility. The platform allows for high throughput measurements in 96-well format and all steps are amendable to automation and scale-up. We believe the use of recombinant antibody technology in combination with SRM MS analysis provides a powerful way to reach sensitivity, specificity and reproducibility as well as the opportunity to build resources for fast on demand implementation of novel assays.
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| 7. |
- Teleman, Johan, et al.
(author)
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Automated quality control system for LC-SRM setups.
- 2013
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In: Journal of Proteomics. - : Elsevier BV. - 1874-3919. ; 95:Online 11 April 2013, s. 77-83
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Journal article (peer-reviewed)abstract
- Selected reaction monitoring (SRM) is emerging as a standard tool for high-throughput protein quantification. For reliable and reproducible SRM protein quantification it is essential that system performance is stable. We present here a quality control workflow that is based on repeated analysis of a standard sample to allow insight into the stability of the key properties of a SRM setup. This is supported by automated software to monitor system performance and display information like signal intensities and retention time stability over time, and alert upon deviations from expected metrics. Utilising the software to evaluate 407 repeated injections of a standard sample during half a year, outliers in relative peptide signal intensities and relative peptide fragment ratios are identified, indicating the need for instrument maintenance. We therefore believe that the software could be a vital and powerful tool for any lab regularly performing SRM, increasing the reliability and quality of the SRM platform. BIOLOGICAL SIGNIFICANCE: Selected reaction monitoring (SRM) mass spectrometry is becoming established as a standard technique for accurate protein quantification. However, to achieve the required quantification reproducibility of the liquid chromatography (LC)-SRM setup, system performance needs to be monitored over time. Here we introduce a workflow with associated software to enable automated monitoring of LC-SRM setups. We believe that usage of the presented concepts will further strengthen the role of SRM as a reliable tool for protein quantification. This article is part of a Special Issue entitled: Standardization and Quality Control.
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| 8. |
- Teleman, Johan, et al.
(author)
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Automated Selected Reaction Monitoring Software for Accurate Label-Free Protein Quantification.
- 2012
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In: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 11:7, s. 3766-3773
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Journal article (peer-reviewed)abstract
- Selected reaction monitoring (SRM) is a mass spectrometry method with documented ability to quantify proteins accurately and reproducibly using labeled reference peptides. However, the use of labeled reference peptides becomes impractical if large numbers of peptides are targeted and when high flexibility is desired when selecting peptides. We have developed a label-free quantitative SRM workflow that relies on a new automated algorithm, Anubis, for accurate peak detection. Anubis efficiently removes interfering signals from contaminating peptides to estimate the true signal of the targeted peptides. We evaluated the algorithm on a published multisite data set and achieved results in line with manual data analysis. In complex peptide mixtures from whole proteome digests of Streptococcus pyogenes we achieved a technical variability across the entire proteome abundance range of 6.5-19.2%, which was considerably below the total variation across biological samples. Our results show that the label-free SRM workflow with automated data analysis is feasible for large-scale biological studies, opening up new possibilities for quantitative proteomics and systems biology.
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| 9. |
- Waldemarson, Sofia, et al.
(author)
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Protein Expression Changes in Ovarian Cancer during the Transition from Benign to Malignant.
- 2012
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In: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 11:5, s. 2876-2889
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Journal article (peer-reviewed)abstract
- Epithelial ovarian carcinoma has in general a poor prognosis since the vast majority of tumors are genomically unstable and clinically highly aggressive. This results in rapid progression of malignancy potential while still asymptomatic and thus in late diagnosis. It is therefore of critical importance to develop methods to diagnose epithelial ovarian carcinoma at its earliest developmental stage, that is, to differentiate between benign tissue and its early malignant transformed counterparts. Here we present a shotgun quantitative proteomic screen of benign and malignant epithelial ovarian tumors using iTRAQ technology with LC-MALDI-TOF/TOF and LC-ESI-QTOF MS/MS. Pathway analysis of the shotgun data pointed to the PI3K/Akt signaling pathway as a significant discriminatory pathway. Selected candidate proteins from the shotgun screen were further confirmed in 51 individual tissue samples of normal, benign, borderline or malignant origin using LC-MRM analysis. The MRM profile demonstrated significant differences between the four groups separating the normal tissue samples from all tumor groups as well as perfectly separating the benign and malignant tumors with a ROC-area of 1. This work demonstrates the utility of using a shotgun approach to filter out a signature of a few proteins only that discriminates between the different sample groups.
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