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Protein Array Protocols Protein Array Protocols	Protein Array Bioinformatics Protein Array bioinformatiku	Learn about Protein Arrays Saznajte više o protein Razvrstati	Protein Array Kits and Products Protein Array Kits i proizvodi	Protein Array Forum Protein Array Forum	Proteomic News Proteomic Novosti

Protein Microarray Capture Molecules and Their Limitations Protein Microarray Capture molekula i njihove granice

Protein and Antibody Microarray Chips Protein i Antitijelo Microarray Chips

Capture Molecules and Their Limitations - Protein Chips Snimite molekula i njihove granice - protein Chips

Capture Molecules and their Limitations Snimite molekula i njihove granice
The most common form of analytical protein arrays are antibody microarrays in which antibodies (or antibody mimics) that bind specific antigens are arrayed on a glass slide at high density. Najčešći oblik analitičke protein polja su antitijela microarrays u kojoj protutijela (ili antitijela mimics) da svežemo određene antigeni su raspoređenih na staklu slajd na visoke gustoće. A lysate is passed over the array and the bound antigen is detected after washing. A lysate je prešlo je u polje i svezana antigen je otkrivena nakon pranje. The biggest challenge with these methods is producing reagents that identify the protein of interest and with high enough specificity in a high-throughput fashion.  Although antibodies are the traditional reagent of choice for detecting proteins in complex mixtures, polyclonal sera are often not specific and are expensive to produce. Najveći izazov s ovih metoda je identificirati da Reagensi za proizvodnju proteina od interesa i sa dovoljno visoku specifičnost u high-fashion propusnost. Protutijela Iako su tradicionalni reagens izbora za detekciju proteina u složenim mješavinama, polyclonal seruma su često nije specifična i preskupa za proizvodnju. Also, the conventional hybridoma method of producing highly specific monoclonal antibodies is time-consuming, laborious and costly (8). Također, hybridoma konvencionalne metode za proizvodnju visoko specifična monoklonska protutijela je dugo trajati, težačkog i skupo (8).

Several studies using antibodies have recently been conducted despite the obstacles in obtaining specific antibodies. Nekoliko studija pomoću protutijela su nedavno provedena unatoč preprekama u dobivanju specifična protutijela.  In one of the largest studies to date, Sreekumar et al. U jednoj od najvećih studija na datum, Sreekumar et al. spotted 146 distinct antibodies on glass to monitor the alternations of protein quantity in LoVo colon carcinoma cells. uočena 146 različita protutijela na staklu da je monitor alternations of protein u količini LoVo Karcinom debelog crijeva stanica. Their results revealed radiation-induced up-regulation of many interesting proteins, including p53, DNA fragmentation factor 40 and 45, tumour necrosis factor-related ligand, as well as down-regulated proteins (58). Njihovi rezultati su pokazale radiation-induced up-regulacija mnoge zanimljive proteina, uključujući i p53, DNA fragmentacije faktor 40 i 45, tumor nekroza faktora vezane uz ligand, kao i down-regulirani proteina (58).

To date, most antibody microarrays were produced with several dozen or a few hundred commercially available poly- or mono-clonal antibodies.  Although tens of thousands of antibodies are commercially available, this number is insufficient because for most proteins there are no available antibodies.  The fact that many antibodies are glycosylated and contain large protein-based supporting structures means that they often cross-react with more than one target protein.  This can contribute to a large number of false positives (9).  Thus another problem has been obtaining high-specificity antibodies. Do danas, većina antitijela microarrays su proizvedene s nekoliko desetak ili nekoliko stotina komercijalno dostupnih poli-ili mono-clonal protutijela. Iako desetaka tisuća protutijela su komercijalno dostupni, ovaj broj je nedostatan, jer za većinu proteina nema dostupnih protutijela. Činjenica da su mnogi protutijela glycosylated i sadrže velike protein-based potpornih struktura znači da su oni često unakrsno reagiraju s više od jedne target protein. To mogu doprinijeti velikom broju neistinit određen (9). Tako je još jedan problem je dobivanja visoko - specifičnost protutijela.

One of the greatest problem with antibody arrays is specificity. Jedan od najvećih problema s polja je specifičnost antitijela. Proteins are often present in a very large dynamic range (106); thus, reagents that might have high affinity for one protein, but are low affinity for another will still exhibit detection of the lower affinity protein if it is much more prevalent (9). Proteini su često prisutni u vrlo veliki dinamički raspon (106); ovako, Reagensi koje bi mogle imati visok afinitet za jedan protein, ali su niski afinitet za neki drugi će i dalje izlagati otkrivanje donjem afinitet proteina ako je puno više prevladava (9) . One group investigated the ability of 115 well-characterized antibody–antigen pairs to react in high-density microarrays on modified glass slides. Jedna skupina istražuje mogućnost od 115 dobro karakterizira antitijelo-antigen parova da reagiraju na visoke gustoće microarrays na izmijenjeni staklo dijapozitiva. 30% of the pairs showed the expected linear relationships, indicating that a fraction of the antibodies were suitable for quantitative analysis (33). 30% od parova pokazala se očekuje linearna veza, navodeci da u djeliću protutijela su pogodne za kvantitativna analiza (33). Many groups have been using sandwich assays to avoid this problem.  A sandwich assay is performed by spotting the first antibody on the array and then detecting the using a second antibody that recognizes a different part of the proteins. Mnoge grupe su pomoću sendvič eseji kako biste izbjegli taj problem. Sendvič esej je, koju provode osobe za otkrivanje prve antitijela na polje, a zatim otkrivanje i koristite drugi antitijelo koje prepoznaje drugu dio od proteina. This approach dramatically increases the specificity of the antigen detection, but required that a least two high-quality antibodies exist for each antigen to be detected (8). Ovakav pristup uvelike povećava specifičnost na antigen za otkrivanje, ali da je potrebno najmanje dva visokokvalitetne protutijela postoje za svaki antigen biti otkrivena (8).

Next: Antibody Microarrays: Problems and Solutions Next: Antitijelo Microarrays: problemi i rješenja

References for Protein and Antibody Microarrays Reference za protein i Antitijelo Microarrays

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Introduction and Background to Protein Chips and Antibody Chips. Uvod i pozadina na protein Chips i Antitijelo Chips.

Types of Antibody and Protein Chips Vrste Antitijelo i protein Chips