Quantification of Aldehyde Functional Groups on Polymeric Microbeads via Catch and Release of Reporters
- Aldehyde-functionalized materials have found broad use in bioconjugation applications. For example, coupling of aldehyde surface groups with to proteins or amine-functionalized oligonucleotides can readily produce biomolecule-covered chip microarray and bead surfaces for multiplex analyses. Additionally, aldehyde-modified nanoparticles can possess bioadhesive properties that can extend their retention time in biological compartments. These emerging novel bioanalytical applications call for reliable tools and methods to detect and quantify accessible aldehyde functionalities. We present here a straightforward concept to quantify the amount of accessible aldehyde moieties on the surface of polymethylmethacrylate (PMMA) particles through the specific binding and subsequent release of small reporter molecules such as absorbing and fluorescent dyes utilizing hydrazone formation as a reversible covalent labeling strategy. Unbound reporter molecules can be easily removed by washing steps,Aldehyde-functionalized materials have found broad use in bioconjugation applications. For example, coupling of aldehyde surface groups with to proteins or amine-functionalized oligonucleotides can readily produce biomolecule-covered chip microarray and bead surfaces for multiplex analyses. Additionally, aldehyde-modified nanoparticles can possess bioadhesive properties that can extend their retention time in biological compartments. These emerging novel bioanalytical applications call for reliable tools and methods to detect and quantify accessible aldehyde functionalities. We present here a straightforward concept to quantify the amount of accessible aldehyde moieties on the surface of polymethylmethacrylate (PMMA) particles through the specific binding and subsequent release of small reporter molecules such as absorbing and fluorescent dyes utilizing hydrazone 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 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 cleavage. 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.…
Autor*innen: | Nithiya Nirmalananthan-Budau |
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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: | 29th Annual Conference of the German Society for Cytometry |
Veranstaltungsort: | Berlin, Germany |
Beginndatum der Veranstaltung: | 25.09.2019 |
Enddatum der Veranstaltung: | 27.09.2019 |
Verfügbarkeit des Dokuments: | Datei im Netzwerk der BAM verfügbar ("Closed Access") |
Datum der Freischaltung: | 28.11.2019 |
Referierte Publikation: | Nein |