Rajan, Malini ; Mortusewicz, Oliver ; Rothbauer, Ulrich ; Hastert, Florian D. ; Schmidthals, Katrin ; Rapp, Alexander ; Leonhardt, Heinrich ; Cardoso, M. Cristina (2015)
Generation of an alpaca-derived nanobody recognizing γ-H2AX.
In: FEBS open bio, 5
Artikel, Bibliographie
Kurzbeschreibung (Abstract)
Post-translational modifications are difficult to visualize in living cells and are conveniently analyzed using antibodies. Single-chain antibody fragments derived from alpacas and called nanobodies can be expressed and bind to the target antigenic sites in living cells. As a proof of concept, we generated and characterized nanobodies against the commonly used biomarker for DNA double strand breaks γ-H2AX. In vitro and in vivo characterization showed the specificity of the γ-H2AX nanobody. Mammalian cells were transfected with fluorescent fusions called chromobodies and DNA breaks induced by laser microirradiation. We found that alternative epitope recognition and masking of the epitope in living cells compromised the chromobody function. These pitfalls should be considered in the future development and screening of intracellular antibody biomarkers.
| Typ des Eintrags: | Artikel |
|---|---|
| Erschienen: | 2015 |
| Autor(en): | Rajan, Malini ; Mortusewicz, Oliver ; Rothbauer, Ulrich ; Hastert, Florian D. ; Schmidthals, Katrin ; Rapp, Alexander ; Leonhardt, Heinrich ; Cardoso, M. Cristina |
| Art des Eintrags: | Bibliographie |
| Titel: | Generation of an alpaca-derived nanobody recognizing γ-H2AX. |
| Sprache: | Englisch |
| Publikationsjahr: | 2015 |
| Titel der Zeitschrift, Zeitung oder Schriftenreihe: | FEBS open bio |
| Jahrgang/Volume einer Zeitschrift: | 5 |
| Kurzbeschreibung (Abstract): | Post-translational modifications are difficult to visualize in living cells and are conveniently analyzed using antibodies. Single-chain antibody fragments derived from alpacas and called nanobodies can be expressed and bind to the target antigenic sites in living cells. As a proof of concept, we generated and characterized nanobodies against the commonly used biomarker for DNA double strand breaks γ-H2AX. In vitro and in vivo characterization showed the specificity of the γ-H2AX nanobody. Mammalian cells were transfected with fluorescent fusions called chromobodies and DNA breaks induced by laser microirradiation. We found that alternative epitope recognition and masking of the epitope in living cells compromised the chromobody function. These pitfalls should be considered in the future development and screening of intracellular antibody biomarkers. |
| Fachbereich(e)/-gebiet(e): | 10 Fachbereich Biologie 10 Fachbereich Biologie > Cell Biology and Epigenetics |
| Hinterlegungsdatum: | 09 Nov 2015 08:06 |
| Letzte Änderung: | 09 Nov 2015 08:06 |
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