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Rapid and simple preparation of thiol-ene emulsion-templated monoliths and their application as enzymatic microreactors.

Lab on a chip (2015-04-09)
Josiane P Lafleur, Silja Senkbeil, Jakub Novotny, Gwenaël Nys, Nanna Bøgelund, Kasper D Rand, Frantisek Foret, Jörg P Kutter
ABSTRACT

A novel, rapid and simple method for the preparation of emulsion-templated monoliths in microfluidic channels based on thiol-ene chemistry is presented. The method allows monolith synthesis and anchoring inside thiol-ene microchannels in a single photoinitiated step. Characterization by scanning electron microscopy showed that the methanol-based emulsion templating process resulted in a network of highly interconnected and regular thiol-ene beads anchored solidly inside thiol-ene microchannels. Surface area measurements indicate that the monoliths are macroporous, with no or little micro- or mesopores. As a demonstration, galactose oxidase and peptide-N-glycosidase F (PNGase F) were immobilized at the surface of the synthesized thiol-ene monoliths via two different mechanisms. First, cysteine groups on the protein surface were used for reversible covalent linkage to free thiol functional groups on the monoliths. Second, covalent linkage was achieved via free primary amino groups on the protein surface by means of thiol-ene click chemistry and l-ascorbic acid linkage. Thus prepared galactose oxidase and PNGase F microreactors demonstrated good enzymatic activity in a galactose assay and the deglycosilation of ribonuclease B, respectively.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
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L-Ascorbic acid, BioUltra, ≥99.5% (RT)
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L-Ascorbic acid, puriss. p.a., ≥99.0% (RT)
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D-(+)-Galactose, meets analytical specification of Ph. Eur., BP
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L-Ascorbic acid, puriss. p.a., ACS reagent, reag. ISO, Ph. Eur., 99.7-100.5% (oxidimetric)
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D-(+)-Galactose, BioXtra, pH 5.0-7.0 (20 °C, 1 M in H2O), ≥99%
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D-(+)-Galactose, ≥99%
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D-(+)-Galactose, powder, anhydrous, BioReagent, suitable for cell culture, suitable for insect cell culture
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D-(+)-Galactose, ≥98%
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Acetylsalicylic acid, analytical standard
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L-Ascorbic acid, BioXtra, ≥99.0%, crystalline
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L-Ascorbic acid, reagent grade, crystalline
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L-Ascorbic acid, suitable for plant cell culture
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L-Ascorbic acid, meets USP testing specifications
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L-Ascorbic acid, suitable for cell culture, suitable for plant cell culture, ≥98%
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L-Ascorbic acid, FCC, FG
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SAFC
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