A Novel Subtractive Antibody Phage Display Method to Discover Disease Markers
| Autor: | Jos M.H. Raats, Hilde E. Roossien, Kalok Cheung, Ger J. M. Pruijn, Danielle Hof |
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| Rok vydání: | 2006 |
| Předmět: |
Cell Extracts
Cytoplasm Phage display Apoptosis Computational biology Biochemistry Antibodies Epitope Autoimmune Diseases Ribonucleoprotein U1 Small Nuclear Analytical Chemistry law.invention Epitopes Jurkat Cells Splicing factor Antigen Peptide Library law Humans Biotinylation Antigens Peptide library Molecular Biology Ribonucleoprotein biology Molecular biology Recombinant Proteins Research Design biology.protein Recombinant DNA Antibody Biomarkers HeLa Cells |
| Zdroj: | Molecular & Cellular Proteomics. 5:245-255 |
| ISSN: | 1535-9476 |
| Popis: | Today's research demands fast identification of potential diagnostic and therapeutic targets. We describe a novel phage display strategy to identify disease-related proteins that are specifically expressed in a certain (diseased) tissue or cells. Phages displaying antibody fragments are selected on complex protein mixtures in a two-step manner combining subtractive selection in solution with further enrichment of specific phages on two-dimensional Western blots. Targets recognized by the resulting recombinant antibodies are immunoaffinity-purified and identified by mass spectrometry. We used antibody fragment libraries from autoimmune patients to discover apoptosis-specific and disease-related targets. One of the three identified targets is the U1-70K protein, a marker for systemic lupus erythematosus overlap disease. Interestingly the epitope on U1-70K recognized by the selected recombinant antibody was shown to be apoptosis-dependent, and such epitopes are believed to be involved in breaking tolerance to self-antigens. The other two proteins were identified as polypyrimidine tract-binding protein-associated splicing factor (PSF)/nuclear RNA- and DNA-binding protein of 54 kDa (p54nrb) and heterogeneous ribonucleoprotein C. |
| Databáze: | OpenAIRE |
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