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. 2022 Mar 10;9:835302. doi: 10.3389/fmolb.2022.835302

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Copyright © 2022 Fagbadebo, Kaiser, Zittlau, Bartlick, Wagner, Froehlich, Jarjour, Nueske, Scholz, Traenkle, Macek and Rothbauer.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Comparison of monovalent and site specifically conjugated bivalent M41- and M114-Nbs in immunofluorescence. For comparable IF analysis HeLa cells transiently expressing GFP-Miro1 (left panel) or wildtype (wt) HeLa cells (right panel) were fixed and stained with the mono- or bivalent Nbs conjugated either chemically (M41_{647_NHS}, M114_{647_NHS}) or site-specifically via sortagging (_bivM41_{647_sort}, _bivM114_{647_sort}) to AlexaFluor 647 (647). As positive control respective cells were stained with an anti-Miro1 IgG followed by detection with a secondary antibody conjugated to AlexaFluor 647 (bottom row). Representative fluorescence images are shown from three independent biological replicates. Nuclei were counterstained with DAPI. Scale bar 20 µm.