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Issue 11, 2016
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Element labeling of antibody fragments for ICP-MS based immunoassays

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Abstract

This work introduces and evaluates the use of recombinantly produced antigen binding fragments (Fab) for ICP-MS based immunoassays. A labeling procedure for a genetically modified anti HER2-specific Fab, so-called ThioFab, with rare earth element labeled macrocycles, and its characterization via liquid chromatography inductively coupled plasma mass spectrometry is presented. The employment of this engineered ThioFab, which carries a recombinantly introduced cysteine allows accurate control of the labeling stoichiometry. The labeling procedure was performed after coordination of selected lanthanides by the complexing DOTA–maleimide-moiety and subsequent linking of the complex to the ThioFab. With the introduced cysteine residue, only a mild reduction step was necessary prior to the attachment of the lanthanide–DOTA–maleimide preventing fragmentation of the Fab-molecule. Pre-optimization steps were performed with human serum albumin as the model protein. By applying the optimized labeling procedure the labeling degree of the genetically modified ThioFab was increased to 90.5 ± 0.5%. The formation of the final ThioFab/antigen conjugate employing Anti-HER2/HER2 as the model system was successfully demonstrated indicating the high potential of the new approach for immunoassays in imaging and flow-cytometry based assays.

Graphical abstract: Element labeling of antibody fragments for ICP-MS based immunoassays

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Publication details

The article was received on 13 Jul 2016, accepted on 06 Oct 2016 and first published on 06 Oct 2016


Article type: Technical Note
DOI: 10.1039/C6JA00252H
Citation: J. Anal. At. Spectrom., 2016,31, 2330-2337
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    Element labeling of antibody fragments for ICP-MS based immunoassays

    T. Mairinger, G. Wozniak-Knopp, F. Rüker, G. Koellensperger and S. Hann, J. Anal. At. Spectrom., 2016, 31, 2330
    DOI: 10.1039/C6JA00252H

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