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Quantification of Aldehydes via Catch and Release of Reporter Chromophores on Polymeric Microbeads

  • Aldehyde-functionalized materials have found widespread use in bioconjugation applications. For example, coupling of aldehyde surface groups with proteins, peptides or amine-functionalized oligonucleotides can readily produce biomolecule-decorated chip and bead surfaces for multiplex analyses. Furthermore, aldehyde-modified nanoparticles can possess bioadhesive properties that can prolong their retention time in biological compartments. These emerging novel bioanalytical and biomedical applications call for reliable tools and methods to detect and quantify accessible aldehyde functionalities on the surface of 2D- and 3D-supports. We present here a versatile concept to quantify the amount of accessible aldehyde moieties on the surface of PMMA particles through the specific binding and subsequent release of small reporter molecules such as absorbing and fluorescent dyes utilizing acylhydrazone formation as a reversible covalent labeling strategy. Unbound reporter molecules can be easilyAldehyde-functionalized materials have found widespread use in bioconjugation applications. For example, coupling of aldehyde surface groups with proteins, peptides or amine-functionalized oligonucleotides can readily produce biomolecule-decorated chip and bead surfaces for multiplex analyses. Furthermore, aldehyde-modified nanoparticles can possess bioadhesive properties that can prolong their retention time in biological compartments. These emerging novel bioanalytical and biomedical applications call for reliable tools and methods to detect and quantify accessible aldehyde functionalities on the surface of 2D- and 3D-supports. We present here a versatile concept to quantify the amount of accessible aldehyde moieties on the surface of PMMA particles through the specific binding and subsequent release of small reporter molecules such as absorbing and fluorescent dyes utilizing acylhydrazone formation as a reversible covalent labeling strategy. Unbound reporter molecules can be easily removed by washing steps, eliminating inaccuracies caused by unspecific adsorption to hydrophobic surfaces. Cleavage of the hydrazones at acidic pH assisted by a carbonyl trap releases the optical reporters rapidly and quantitatively and allows for their optical detection at low concentration. Importantly, this strategy separates the signal-generating molecules from the bead surface, thereby circumventing common pitfalls of optical assays associated with light scattering and signal distortions due to binding-induced changes in reporter fluorescence and quenching dye-dye interactions on crowded material surfaces. The potential of this catch-and-release strategy for surface group quantification is representatively demonstrated for a set of microparticles functionalized with different aldehyde densities. This concept is validated by a colorimetric assay with a different optical probe, which contains a reductively cleavable disulfide bond and a reporter that can be quantified photometrically in solution after its release. The excellent match of the results of both optical assays confirms their suitability for the rapid and sensitive quantification of aldehydes on microbead surfaces. These simple catch-and-release assays are excellent tools for process control during bead fabrication and the comparison of different bead batches. Their potential for predicting biomolecule coupling efficiencies in bioconjugation reactions is currently assessed in reductive amination reactions with streptavidin.zeige mehrzeige weniger

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Metadaten
Autor*innen:Nithiya Nirmalananthan-Budau
Koautor*innen:Alexander Roloff, Bastian Rühle, H. Borcherding, T. Thiele, U. Schedler, Ute Resch-Genger
Dokumenttyp:Posterpräsentation
Veröffentlichungsform:Präsentation
Sprache:Englisch
Jahr der Erstveröffentlichung:2019
Organisationseinheit der BAM:1 Analytische Chemie; Referenzmaterialien
1 Analytische Chemie; Referenzmaterialien / 1.2 Biophotonik
DDC-Klassifikation:Naturwissenschaften und Mathematik / Chemie / Analytische Chemie
Freie Schlagwörter:CHO functions; Catch and release assay; Optical quantification; Polymer particles
Themenfelder/Aktivitätsfelder der BAM:Chemie und Prozesstechnik
Veranstaltung:Tag der Chemie
Veranstaltungsort:Berlin, Germany
Beginndatum der Veranstaltung:11.07.2019
Verfügbarkeit des Dokuments:Datei im Netzwerk der BAM verfügbar ("Closed Access")
Datum der Freischaltung:28.11.2019
Referierte Publikation:Nein
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