Isoenzyme-directed selection and characterization of anti-creatine kinase single chain Fv antibodies from a human phage display library. - Archive ouverte HAL Access content directly
Journal Articles Biochimica et Biophysica Acta - Molecular Cell Research Year : 2002

Isoenzyme-directed selection and characterization of anti-creatine kinase single chain Fv antibodies from a human phage display library.

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Abstract

Epitopes differing among isoenzymes of creatine kinase (CK) are apparently limited in number and poorly immunogenic in vivo. Especially for the BB-CK isoenzyme, very few monoclonal antibodies (mAb) are available. Here, we use in vitro selection with a synthetic human phage display antibody library and develop isoenzyme competition and peptide panning strategies to obtain human single chain Fv (scFv) antibodies against specific CK isoenzymes. We isolated and characterized seven scFv clones that recognize native as well as denatured cytosolic BB-CK in ELISA, immunoblot, immunofluorescence histochemistry and surface plasmon resonance (SPR) spectroscopy. To a variable but minor degree, they also react with cytosolic MM-CK, but not with mitochondrial CK isoenzymes. Epitope mapping revealed that the scFv antibodies recognize different BB-CK epitopes, including the N-terminus and the isoenzyme-specific box, a highly conserved sequence of unknown function for which no mAb were available so far. With a K(D) of 3.5-9.6 x 10(-7) M, the isolated scFv compare favorably with mouse mAb and may overcome certain of their limitations. Our results demonstrate the advantages of in vitro antibody selection for the generation of isoenzyme-specific antibodies.
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Dates and versions

inserm-00390848 , version 1 (02-06-2009)

Identifiers

  • HAL Id : inserm-00390848 , version 1
  • PUBMED : 12427547

Cite

Uwe Schlattner, Christof Reinhart, Thorsten Hornemann, Malgorzata Tokarska-Schlattner, Theo Wallimann. Isoenzyme-directed selection and characterization of anti-creatine kinase single chain Fv antibodies from a human phage display library.. Biochimica et Biophysica Acta - Molecular Cell Research, 2002, 1579 (2-3), pp.124-32. ⟨inserm-00390848⟩

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