| 231. |
- Sincic, Viktor, et al.
(författare)
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Transcriptomic profiling of T-cell populations in non-muscle invasive and muscle invasive bladder cancer.
- 2020
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Ingår i: Journal for ImmunoTherapy of Cancer. - 2051-1426. ; 8:Suppl. 3
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Konferensbidrag (refereegranskat)abstract
- Background: Bladder cancer is categorized as non-muscle invasive (NMIBC) or muscle invasive (MIBC). NIMBC makes up around 70% of the cases and although it is less aggressive, the recurrence rate is 50-70%, thus requiring extensive monitoring. Additionally, there is a risk of progression into MIBC with a 5-year survival of only 50% even when treated with radical cystectomy. Immune checkpoint inhibitors have shown promising results for treatment of bladder cancer; however, only around 30% of patients have a therapeutic effect and novel therapies are thus required. With the aim of pinpointing novel targets for T-cell based therapy, we have performed transcriptomic profiling of specific T cell populations in MIBC and NMIBC, as well as in control bladder tissue.Methods: Muscle-invasive (n=7) as well as non-muscle invasive (n=13) bladder tumor biopsies were obtained from untreated patients and control bladder tissue (n=7). Upon digestion, cells were stained with an antibody panel to enable sorting of CD8+ cytotoxic T-cells (CD8T), CD4+ T-helper cells (Th) and regulatory T-cells (Treg) using fluorescence activated cell sorting. RNA was extracted and subject to sequencing. Differential gene expression analysis was performed, using DESeq2 (genes with padjResults: Principal component analysis demonstrated that CD8T, unlike Th and Tregs, cluster according to the invasiveness of the disease. Accordingly, many genes were significantly differentially expressed between CD8T in MIBC and NMIBC compared to control, and also between CD8T in MIBC compared to NMIBC. Several genes associated with CD8 T-cell exhaustion were significantly upregulated in MIBC compared to both NMIBC and control. Further, GSEA results indicated biological differences of the CD8T compartment between different tumor stages.Conclusion: The gene expression profiles of CD8 T-cells were significantly different in NMIBC, MIBC and control. The transcriptional profiles give clues on biological differences and disease progression and can be relevant for development of novel treatment strategies.
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| 232. |
- Sjogren-Jansson, Eva, et al.
(författare)
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Production of human monoclonal antibodies in dialysis tubing
- 1991
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Ingår i: Hybridoma. - : Mary Ann Liebert Inc. - 0272-457X. ; 10:3, s. 411-419
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Tidskriftsartikel (refereegranskat)abstract
- Human x mouse hybridoma cells were grown in dialysis tubing (DT) to obtain large quantities of human monoclonal antibodies (MAb). Hybridoma cells were grown inside the DT, which was placed in a tissue culture flask containing medium. The medium inside the DT was supplemented with different additives which may be selected depending on the intended use of the MAb. About 10-50 times higher concentrations of immunoglobulin (Ig) were obtained after cultivation in DT compared to conventional tissue culture (CTC) for 2 days. The purity of the MAb was high which facilitated further antibody purification. Production of human MAb in DT proved to be excellent for evaluation studies in laboratory scale. It does not require expensive equipment and several hybridomas can be grown simultaneously.
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| 233. |
- Skoog, Petter, et al.
(författare)
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Tumor tissue protein signatures reflect histological grade of breast cancer
- 2017
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203.
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Tidskriftsartikel (refereegranskat)abstract
- Histological grade is one of the most commonly used prognostic factors for patients diagnosed with breast cancer. However, conventional grading has proven technically challenging, and up to 60% of the tumors are classified as histological grade 2, which represents a heterogeneous cohort less informative for clinical decision making. In an attempt to study and extend the molecular puzzle of histologically graded breast cancer, we have in this pilot project searched for additional protein biomarkers in a new space of the proteome. To this end, we have for the first time performed protein expression profiling of breast cancer tumor tissue, using recombinant antibody microarrays, targeting mainly immunoregulatory proteins. Thus, we have explored the immune system as a disease-specific sensor (clinical immunoproteomics). Uniquely, the results showed that several biologically relevant proteins reflecting histological grade could be delineated. In more detail, the tentative biomarker panels could be used to i) build a candidate model classifying grade 1 vs. grade 3 tumors, ii) demonstrate the molecular heterogeneity among grade 2 tumors, and iii) potentially re-classify several of the grade 2 tumors to more like grade 1 or grade 3 tumors. This could, in the long-term run, lead to improved prognosis, by which the patients could benefit from improved tailored care
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| 234. |
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| 235. |
- Steinhauer, Cornelia, et al.
(författare)
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Biocompatability of surfaces for antibody microarrays: Design of macroporous silicon substrates
- 2005
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Ingår i: Micro Total Analysis Systems 2004, Vol 2. - 0260-6291. ; :297, s. 121-123
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Konferensbidrag (refereegranskat)abstract
- Antibody microarray is a novel technology with great promise within proteomics. Intense work is under way to evolve this methodology into the high-throughput proteomic research tool needed by the research community. Despite recent advances, there is a growing need for additional highperformance substrates for antibody microarrays as well as for protein arrays in general. In this study, we have sucessfully designed novel, highly biocompatible and well-performing silicon-based supports that has the capacity to play a significant role within current and future antibody and protein microarray applications within the field of proteomics.
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| 236. |
- Steinhauer, Cornelia, et al.
(författare)
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Biocompatibility of surfaces for antibody microarrays: design of macroporous silicon substrates
- 2005
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Ingår i: Analytical Biochemistry. - : Elsevier BV. - 1096-0309 .- 0003-2697. ; 341:2, s. 204-213
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Tidskriftsartikel (refereegranskat)abstract
- Major efforts to develop antibody microarray technology to enable global proteome analysis to be performed in a facile manner are under way. In this process, the design and the properties of the substrate will play crucial roles. In the present study, we have developed novel, highly biocompatible solid supports for microarrays, using adsorbed recombinant human single-framework antibody fragments as probes. Several silicon-based supports, including planar silicon, micro- and macroporous silicon, and nitrocellulose-coated variants thereof, were designed and evaluated in a stepwise procedure. The surfaces were scored based on biocompatibility and probe binding capacity as judged by spot morphology, signal intensities, signal to noise ratios, dynamic range, sensitivity, and reproducibility. A set of five commercially available substrates, selected to represent a set of supports providing different surface and coupling chemistries, was used as reference surfaces. The results showed that several well-performing silicon-based supports could be designed; in particular, a nitrocellulose-coated macroporous variant, MAP3-NC7, received the highest scores. In comparison, MAP3-NC7 displayed properties equal to or better than those of the reference substrates. Taken together, designed surfaces based on silicon can undoubtedly meet the requirements of the next generation of solid supports for antibody microarrays. (c) 2004 Elsevier Inc. All rights reserved.
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| 237. |
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| 238. |
- Steinhauer, Cornelia, et al.
(författare)
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Improved affinity coupling for antibody microarrays: Engineering of double-(His)(6)-tagged single framework recombinant antibody fragments
- 2006
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Ingår i: Proteomics. - : Wiley. - 1615-9861 .- 1615-9853. ; 6:15, s. 4227-4234
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Tidskriftsartikel (refereegranskat)abstract
- Antibody-based microarray is a novel technology with great promise in biomedicine that will provide unique means to perform global proteome analysis. In the process of designing the high-density antibody microarrays required, several critical key issues have been identified that remain to be resolved. In particular, there is a great need for specific and selective approaches enabling non-purified probes to be directly purified, orientated and coupled in a generic one-step procedure directly on the chip. In this study, we report on the successful design of affinity-tagged human recombinant single-chain fragment variable antibody fragments for improved affinity coupling in array applications. By replacing the standard single-histidine (His)(6)-tag with a consecutive double-(His)(6)-tag, the binding to Ni2+-nitrilotriacetic acid-coated substrates was significantly improved. Surface plasmon resonance analysis showed a significantly tighter binding with at least a threefold slower dissociation. The improved binding characteristics thus enabled non-purified probes even in the format of crude expression supernatants to be directly applied thereby eliminating the need for any time-consuming pre-purification step(s) prior to the immobilization. While the double-(HiS)(6)-tag probes were found to be expressed equally well as compared to the single-(His)(6)-tag probes, they displayed better long-term functional on-chip stability. Taken together, the results demonstrate the generic potential of double-(HiS)(6)-tag recombinant antibodies for the facile fabrication of high-density antibody microarrays.
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| 239. |
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| 240. |
- Steinhauer, Cornelia, et al.
(författare)
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Single framework recombinant antibody fragments designed for protein chip applications
- 2002
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Ingår i: BioTechniques. - 0736-6205. ; :Suppl., s. 38-38
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Tidskriftsartikel (refereegranskat)abstract
- High-throughput proteomics, based on the microarray platform, requires stable, highly functional components that will yield a highly sensitive read-out of low, abundance protein. Although antibodies are the best characterized binding molecules for this purpose, only a fraction of them appear to behave satisfactorily in the chip format. Therefore, high demands need to be placed on their molecular design. In the present study, we have focused an recombinant antibody design based on a single framework for protein chip applications, aiming at defining crucial molecular probe parameters. Our results show that engineered human recombinant scFv antibody fragment, that displayed appropriate biophysical properties (molecular [functional] stability in particular) can be generated, making them prime candidates for high-density antibody arrays. In fact a superior framework that displays both multifaceted adsorption properties and very high functional stability over several months on chips (stored in a dried-out state) was identified Taken together designed scFv fragments based on a single molecular scaffold, readily accessible in Large phage display libraries, can undoubtedly meet the requirements of probe content in antibody microarrays, particularly for global proteome analysis.
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