Subsequent to this exhibition of recent suitable proteomics studies, the participants of the 9th HUPO BPP workshop on Barbados re-defined the HUPO BPP mission. They agreed that:
· the HUPO BPP aims at advancing knowledge and understanding neurodiseases and aging with the purpose to identify prognostic and diagnostic biomarkers as well as to push new diagnostic approaches and medications (e.g. for therapy control)
· the HUPO BPP will perform quantitative proteomics on disease-related brain areas and bodily fluids
· participants will analyze the brain proteome of human as well as mouse models in healthy, diseased and aged status with focus on Alzheimer's and Parkinson's Disease
· the groups will exchange knowledge and data with other HUPO projects and national / international initiatives in the neuroproteomic field, making neuroproteomic research and its results available to the scientific community, especially to the members of the HUPO Project.
With this mission, the HUPO BPP is integrated well in HUPO. Since the HUPO BPP is searching for autoimmune antibodies, the consortium is especially interested in getting access to well characterized monoclonal antibodies and their respective purified antigens in order to elucidate the progression e.g. of Parkinson’s and Alzheimer’s Disease.
Concerning the criteria that have to be fulfilled in successful proteomics it was stated that
· methods have to be robust, so that e.g. the same system can be run several times
· methods have to show a high sensibility, so that e.g. low abundant proteins can be detected
· methods should be available for reasonable costs (for academic institutes ) allowing independent repetitions
· each laboratory has to show the reproducibility of every result. This leads to the consequence that high performance has to be estimated higher than high throughput.
In regard to reagents, technologies and improvements that should be aimed at it was agreed upon the following:
· Reagents: Antibody and protein array development is regarded as important in order to generate trustable tools for identification, validation etc. Reagents should be reasonably priced, e.g. isotopic labelling kits, allowing independent repetitions.
· 2D Poly Acrylamide Gel Electrophoresis: The analytical software has to be improved; the Pinnacle software introduced above could be an alternative. In addition, especially cheaper DIGE dyes are recommended (see also point reagents).
· Capillary Electrophoresis: CE was regarded to be reserved for higher resolution and special applications, e.g. quality profiling.
· Mass Spectrometry in general: Concerning a HUPO BPP-conform mass spectrometry, the dynamic range, the quantification, the robustness, the reproducibility and the sensitivity should be improved. High end instruments should be more reasonably priced.
· nano Liquid Chromatography Mass Spectrometry: To allow reproducible studies, the whole LC system should be inert/non-reactive and oxygen-free. It is important to improve quantification, reproducibility, and robustness to analyse many biological samples in a row.
· Imaging Mass Spectrometry: At the moment 1 order of magnitude is available. Therefore, an improved spatial resolution with a higher dynamic range, improved robustness, improved sensitivity, and improved reproducibility is necessary.
Samples, handling and models have been discussed as follows:
The best model for human is human, so that human tissues are to be preferred (non-affected and affected gene carriers). As of course the sources are limited, bodily fluids should be used, namely CSF and serum. Epilepsy surgeries might be a source, too. Post mortem material is suitable best for special applications as e.g. marker validation, while stem cells were excluded due to EU/US laws and scientific reasons.
Rodent models (transgenic and KO) and cell lines should be analysed for functional and developmental studies. Models will be defined partly according to the HUPO project.
Concerning sample acquisition and handling (storage etc.) it was stressed that standardisation of conditions (esp. for post mortem material) and SOPs are crucial for any study. These guidelines have already been adopted from BrainNet Europe, the Competence Net Dementia, the HUPO PSI as well as the EU-project Proteomics Data Collection ProDaC. These efforts should be forced by grant agencies and journals to achieve a fast and wide acceptance. All co-workers have to be trained very well according to these SOPs to obtain maximum reproducibility. Sample processing, e.g. via cell sorting using specific antibodies, subcellular fractionation, etc, as well as protein extraction and separation/fractionation were also discussed and should be handled similar.
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