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Mischak, Harald; Schiffer, Eric; Zürbig, Petra; Dakna, Mohammed; Metzger, Jochen
Journal of Medical Biochemistry, 10/2009, Volume: 28, Issue: 4Journal Article
Proteome analysis has emerged as a powerful tool to decipher (patho) physiological processes, resulting in the establishment of the field of clinical proteomics. One of the main goals is to discover biomarkers for diseases from tissues and body fluids. Due to the enormous complexity of the proteome, a separation step is required for mass spectrometry (MS)-based proteome analysis. In this review, the advantages and limitations of protein separation by two-dimensional gel electrophoresis, liquid chromatography, surface-enhanced laser desorption/ionization and capillary electrophoresis (CE) for proteomic analysis are described, focusing on CE-MS. CE-MS enables separation and detection of the small molecular weight proteome in biological fluids with high reproducibility and accuracy in one single processing step and in a short time. As sensitive and specific single biomarkers generally may not exist, a strategy to overcome this diagnostic void is shifting from single analyte detection to simultaneous analysis of multiple analytes that together form a disease-specific pattern. Such approaches, however, are accompanied with additional challenges, which we will outline in this review. Besides the choice of adequate technological platforms, a high level of standardization of proteomic measurements and data processing is also necessary to establish proteomic profiling. In this regard, demands concerning study design, choice of specimens, sample preparation, proteomic data mining, and clinical evaluation should be considered before performing a proteomic study. Analiza proteoma je postala je moćno sredstvo za dešifrovanje (pato)fizioloških procesa, što je za rezultat imalo uspostavljanje oblasti kliničke proteomike. Jedan od glavnih ciljeva je otkrivanje biomarkera oboljenja iz tkiva i telesnih tečnosti. Zbog ogromne složenosti proteoma, pri proteomskoj analizi zasnovanoj na masenoj spektrometriji potrebno je izvršiti separaciju. U radu su opisane prednosti i ograničenja proteomske analize pri separaciji proteina putem dvodimenzionalne gel elektroforeze, tečne hromatografije, SELDI i kapilarne elektroforeze (KE), sa fokusom na KE-MS. KE-MS omogućava separaciju i detekciju proteoma male molekularne težine u biološkim tečnostima uz visoku reproducibilnost i preciznost u samo jednom koraku radnog postupka i za kratko vreme. Pošto pojedinačni senzitivni i specifični biomarkeri možda i ne postoje, strategija za premošćivanje te dijagnostičke praznine pomera se sa detekcije pojedinačnog analita na simultanu analizu više analita koji zajedno čine obrazac specifičan za dato oboljenje. Takvi pristupi, međutim, nose sa sobom dodatne izazove, koje ćemo predstaviti u ovom radu. Pored izbora odgovarajućih tehnoloških platformi, neophodan je visok nivo standardizacije proteomskih merenja i obrade podataka kako bi se vršilo profilisanje proteoma. U tom pogledu, zahtevi koji se tiču nacrta studija, izbora primeraka uzoraka, analize proteomskih podataka i kliničke evaluacije trebalo bi da budu razmotreni pre izvođenja proteomske studije.
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