Coeliac disease is a chronic enteropathy triggered by gluten. Patients produce antibodies to gliadin and the autoantigen tissue transglutaminase (tTG). These anti-tTG autoantibodies are disease specific and used in diagnosis. The autoantibodies can be detected by immunofluorescence (the endomysial antibody tests) or by ELISA using recombinant tTG. In vivo tTG associates with fibronectin, which may account for the greater sensitivity of the endomysial antibody assay compared to the ELISA. This project had two aims: to determine whether GST-tagged tTG bound fibronectin and then, using the fibronectin bound tTG, whether a two-tiered ELISA increased anti-tTG binding in coeliac disease patients. First fibronectin was coupled to a solid support and then incubated with tTG. This was then analysed using SDS-PAGE. Secondly, an ELISA with a two-tiered antigen coating was created by coupling tTG to Fn. This mimics the in vivo orientation of the antigen and could theoretically increase anti-tTG binding. Comparative ELISAs were then run to see if anti-tTG binding differed between tTG and fibronectin-coupled tTG antigen coatings. Results showed GST-tagged tTG bound fibronectin. Coupling of tTG to fibronectin gave no improved binding of anti-tTG. On the contrary, most patients tested had decreased anti-tTG binding compared to the normal tTG based ELISA.
| Identifer | oai:union.ndltd.org:UPSALLA1/oai:DiVA.org:uu-155482 |
| Date | January 2011 |
| Creators | Svanqvist, Anna |
| Source Sets | DiVA Archive at Upsalla University |
| Language | English |
| Detected Language | English |
| Type | Student thesis, info:eu-repo/semantics/bachelorThesis, text |
| Format | application/pdf |
| Rights | info:eu-repo/semantics/openAccess |
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