ArticleHepatology research : the official journal of the Japan Society of Hepatology2012
Generation of a cholangiocyte-specific cDNA expression library for the identification of B and T cell autoantigens in murine biliary disease.
Article in Hepatology research : the official journal of the Japan Society of Hepatology, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
aimSeveral mouse models of inflammatory cholangiopathies exist, including biliary atresia, primary biliary cirrhosis, autoimmune hepatitis, and primary sclerosing cholangitis. In an ongoing effort to identify the target antigens of both infiltrating autoreactive T cells and serum autoantibodies, we aimed to generate a cholangiocyte-derived cDNA library capable of expressing a wide variety of proteins.
methodsmRNA was isolated from a normal mouse cholangiocyte cell line and reverse transcribed into cDNA. After initial cloning of the cDNA into a transfer vector (pDONR222), the entire library was shuttled into an Escherichia coli expression vector (pDEST160).
resultsThe library contains 2.3 × 10(6) independent clones and expresses proteins up to 100 kD in molecular weight. Using a variety of techniques, including western blot analysis, mass spectrometry of individual clones, and direct DNA sequencing of plasmids, a number of both ubiquitously expressed and cholangiocyte-specific proteins (e.g. cytokeratin 19) have been identified within.
conclusionA comprehensive mouse cholangiocyte cDNA expression library has been generated and is available for use as a source of multiple cholangiocyte-specific antigens for immunological studies. The library can be used to screen for specificity of T cell lines or hybridomas. Furthermore, this library has potential uses in SEREX analysis of autoantibody reactivity. The cholangiocyte-specific cDNA library is a powerful tool for the identification of target antigens in murine inflammatory cholangiopathies and is available as a shared resource.
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