Uppsala group organization
Dr. Cecilia Lindskog Bergström PI and Site Director HPA Uppsala
Prof. Fredrik Pontén Clinical Director
Jonas Gustavsson Project coordinator
Head: Dr. Cecilia Lindskog Bergström
Anna Maria Clementz (research engineer), Dr. Andreas Digre (research engineer), Dr. Åsa Edvinsson (post doctoral), Jonas Gustavsson (project coordinator), Neda Hekmati (research engineer), Dennis Kesti (biomedical analyst), Borbala Katona (research engineer), Emil Lindström (research engineer), Dr. Loren Méar (post doctoral), Feria Hikmet Noraddin (PhD student), Leo Nore (research engineer), IngMarie Olsson (research engineer), Rutger Schutten (research engineer), Jimmy Vuu (research engineer), Fredrika Hagberg (assistant), Jacob Wakter (biomedical analyst student)
Generation of antibody-based protein data for the Tissue Atlas and the Pathology Atlas:
- Handling and processing of tissues (biobank material)
- Handling and storage of antisera
- Testing of antibodies
- Immunohistochemical staining of tissues slides with approved antibodies
- Scanning of tissue slides and image processing
- Validation of antibody target specificity
- Annotation and final approval of immunohistochemically stained tissues
- Determination of antibody reliability based on enhanced antibody validation strategies
- Generation of knowledge-based protein expression profiles
- Prepare the data for release on the Tissue Atlas and Pathology Atlas
Formalin fixed, paraffin embedded tissue specimens are collected from the Department of Clinical Pathology and Cytology, Uppsala University Hospital and subsequently processed for production of tissue microarrays (TMAs), and protein extraction for Western blot analysis. In addition to standard TMAs, additional tissue samples are used for extended profiling of certain proteins.
Immunohistochemistry is performed using standardized procedures and includes testing antibodies on TMAs and whole sections of tissues in order to create protein expression profiles. All procedures follow strict guidelines and include quality control. Optimal antibody dilution and target specificity is assessed by microscopic examination and comparison of immunohistochemical staining with mRNA data and internal and external gene/protein characterization data. For each approved antibody, a final immunostaining protocol is defined and subsequently applied to the standard 8 full-scale TMAs. For extended profiling of certain proteins, also slides containing extended samples are used.
Approved immunostained slides are scanned to generate high-resolution digital images, using a 20x objective.
Images are imported to an in-house built software to facilitate subsequent manual annotation of images, as well as a subsequent step of curation by an independent second observer.
The generated protein profiles then undergo further evaluation and antibodies that pass quality criteria are assigned a reliability score, a knowledge-based protein expression profile and are scheduled for publication in the next version of the Human Protein Atlas website (www.proteinatlas.org).
Head: Prof Fredrik Pontén
Neda Hekmati (research engineer)
- Develop strategies to identify potential biomarkers based on the HPA database and other efforts.
- Validate proteins that can be used as clinical biomarkers for disease.
- Participate in clinical studies, collect tumor material and clinical data to generate specific cancer TMAs coupled to clinical databases.
- Perform statistical analysis and validate the clinical usefulness of identified biomarkers.
The HPA database is actively mined for potential biomarkers with the aim to identify protein expression patterns that could be of medical or biological significance. The biomarker discovery and validation efforts include both internal projects and external collaborative projects, and include various forms of human disease with an emphasis on cancer. Most projects are focused on identification and validation of biomarker candidates that can fulfill currently unmet clinical needs related to diagnostics, prognostics and treatment prediction.
To pursue such projects, patient cohorts, representing different cancers, are defined and tumor material as well as clinical data is collected. Specifically designed cancer TMAs are produced and used for extended analysis of protein expression patterns to test and validate candidate proteins as useful biomarkers.