This paper reports on the surface modification of plastic microfluidic channels to prepare different biomolecule micropatterns using ultraviolet (UV) photografting methods. The linkage chemistry is based upon UV photopolymerization of acryl monomers to generate thin films (0.01-6 microm) chemically linked to the organic backbone of the plastic surface. The commodity thermoplastic, cyclic olefin copolymer (COC) was selected to build microfluidic chips because of its significant UV transparency and easiness for microfabrication by molding techniques. Once the polyacrylic films were grafted on the COC surface using photomasks, micropatterns of proteins, DNA, and biotinlated conjugates were readily obtained by surface chemical reactions in one or two subsequent steps. The thickness of the photografted films can be tuned from several nanometers up to several micrometers, depending on the reaction conditions. The micropatterned films can be prepared inside the microfluidic channel (on-chip) or on open COC surfaces (off-chip) with densities of functional groups about 10(-7) mol/cm2. Characterization of these films was performed by attenuated-total-reflectance IR spectroscopy, fluorescence microscopy, profilometry, atomic force microscopy, and electrokinetic methods.

The oxidation of glutathione (GSH) by the electrogenerated mediator [IrCl 6 ] 2in the presence of bases (B) to produce the radical GS • , [IrCl 6 ] 3and BH + , was studied by cyclic voltammetry in buffered and unbuffered solution using glassy carbon electrodes. This proton-coupled electron transfer (PCET) is the first step in the oxidation of GSH to produce the disulfide GSSG. The reaction exhibits a slight acceleration of the rate when the phosphate buffer (PB) concentration is raised at constant pH (5.0 or 7.0). The evidence indicates that proton acceptors whose conjugate acids have a pK a lower than GSH (8.7) but higher than GSH •+ can catalyze the reaction even though direct deprotonation of GSH is thermodynamically unfavorable. The value of k obs at 0.005 M of PB pH = 7.0 was equal to 4.3 ± 0.7 × 10 4 M −1 s −1 whereas at 0.200 M, was 2.56 ± 0.03 × 10 5 M −1 s −1 . A kinetic isotope effect (KIE) dependent on [PB] was observed at pD 7.0, confirming that the S−H bond breaks during the rate-determining step. The predominance of a concerted PCET is postulated based on the compliance with the libido rule of general base catalysis, the KIE observed and the finding that stepwise pathways occur in lesser degree. Digital simulations were used to evaluate this mechanism.

T his is a Reply to the Comment by Stanbury et al. 1 on our report about the buffer catalysis observed when glutathione (GSH) is oxidized by electrogenerated IrCl 6 2−

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