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Key Factors for Stable Retention of Fluorophores and Labeled Biomolecules in Droplet-Based Microfluidics
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posted on 2015-02-17, 00:00 authored by Jan-Willi Janiesch, Marian Weiss, Gerri Kannenberg, Jonathon Hannabuss, Thomas Surrey, Ilia Platzman, Joachim P. SpatzWater-in-oil emulsion droplets created
in droplet-based microfluidic
devices have been tested and used recently as well-defined picoliter-sized
3D compartments for various biochemical and biomedical applications.
In many of these applications, fluorescence measurements are applied
to reveal the protein content, spatial distribution, and dynamics
in the droplets. However, emulsion droplets do not always provide
entirely sealed compartments, and partitioning of dyes or labeled
molecules to the oil phase is frequently observed. Therefore, stable
molecular retention in the droplets represents a challenge, and many
physical and chemical key factors of microfluidic system components
have to be considered. In this study, we investigated the retention
of 12 commonly used water-soluble dyes in droplets having six different
aqueous phase conditions. We demonstrate that the physicochemical
properties of the dyes have a major influence on the retention level.
In particular, hydrophilicity has a strong influence on retention,
with highly hydrophilic dyes (LogD < −7) showing stable,
buffer/medium independent retention. In the case of less hydrophilic
dyes, we showed that retention can be improved by adjusting the surfactants
physical properties, such as geometry, length, and concentration.
Furthermore, we analyzed the retention stability of labeled biomolecules
such as antibodies, streptavidin, and tubulin proteins and showed
that stable retention can be strongly dependent on dye and surfactants
selection.