title: Real-time stability testing of air-dried primers and fluorogenic hydrolysis probes stabilized by trehalose and xanthan.
abstract: A method for conserving primers and differently labeled fluorogenic hydrolysis (i.e., TaqMan) probes at ambient conditions is presented. Primers and hydrolysis probes with four different fluorophore-quencher combinations (6- FAM-BHQ1, HEX-BHQ1, ROX-BHQ650, and Cy5-BHQ2) were mixed with trehalose and xanthan at final concentrations of 56 mM and 2.78 mM, respectively. Mixtures were air-dried at 23°C for 30 min on strips composed of cyclo olefin polymer (COP), a material widely used in the manufacturing of in vitro diagnostic (IVD) test carriers. After one year of storage, the functionality of the primers and fluorophore-quencher combinations was validated by real-time polymerase chain reaction (real-time PCR), confirming their stability when stored in the presence of stabilizers, with the best results achieved using trehalose. This approach could be of great benefit for manufacturing IVD systems, for example, for genotyping applications based on multiplexing using different fluorescent dyes.
authors.affiliation: HSG-IMIT-Institut für Mikro- und Informationstechnik, Freiburg, Germany; Laboratory for MEMS Applications, Department of Microsystems Engineering-IMTEK, University of Freiburg, Freiburg, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Rombach
authors.affiliation: HSG-IMIT-Institut für Mikro- und Informationstechnik, Freiburg, Germany.
authors.forename: Dominique
authors.initials: D
authors.lastname: Kosse
authors.affiliation: Laboratory for MEMS Applications, Department of Microsystems Engineering-IMTEK, University of Freiburg, Freiburg, Germany.
authors.forename: Bernd
authors.initials: B
authors.lastname: Faltin
authors.affiliation: BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Wadle
authors.affiliation: HSG-IMIT-Institut für Mikro- und Informationstechnik, Freiburg, Germany; Laboratory for MEMS Applications, Department of Microsystems Engineering-IMTEK, University of Freiburg, Freiburg, Germany; BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany.
authors.forename: Günter
authors.initials: G
authors.lastname: Roth
authors.affiliation: HSG-IMIT-Institut für Mikro- und Informationstechnik, Freiburg, Germany; Laboratory for MEMS Applications, Department of Microsystems Engineering-IMTEK, University of Freiburg, Freiburg, Germany; BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany.
authors.forename: Roland
authors.initials: R
authors.lastname: Zengerle
authors.affiliation: HSG-IMIT-Institut für Mikro- und Informationstechnik, Freiburg, Germany; Laboratory for MEMS Applications, Department of Microsystems Engineering-IMTEK, University of Freiburg, Freiburg, Germany.
authors.forename: Felix
authors.initials: F
authors.lastname: von Stetten
mesh_terms: D017931:DNA Primers / Q000737:chemistry*; D005079:Excipients / Q000737:chemistry*; D005456:Fluorescent Dyes; D006868:Hydrolysis; D011135:Polysaccharides, Bacterial / Q000737:chemistry*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D014199:Trehalose / Q000737:chemistry*
chemical_list: D017931:DNA Primers; D005079:Excipients; D005456:Fluorescent Dyes; D011135:Polysaccharides, Bacterial; D014199:Trehalose; C002563:xanthan gum
keywords: air-drying; preservation; primer; probe; stability testing; trehalose; xanthan