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Sensors 2020, 20, 1333 12 of 15 blocked surfaces. This demonstrates the principal suitability of IPEs for DNA-sensing applications and the used conditions with amplification buffer should allow a facile adaption for real applications. As the results indicate that the higher number of porous layers does not contribute to the hybridization signal, the use of 3-layered electrodes is advantageous in terms of a reduced capacitive background and a fast and resource-efficient production process. Compared to the tested SPEs, an activation step (CV in H2SO4) prior to functionalization was not necessary. The results for the 2.5D chip (SI Figure S11) successfully demonstrate the integration of electrodes in the fluidic chip over the sidewalls of the chamber, if they feature a slope of 45◦ and no sharp edges. This enables the facile assembly of electrochemical lab-on-a-chip systems that can be sealed with comparably simple methods like pressure-sensitive adhesives. The commonly used approach [2,44] of bonding a separate fluidic layer to the planar electrode layer becomes then obsolete. 5. Conclusions A commercially available nanoparticle gold ink was deposited with a drop-on-demand inkjet-printer on COC. Because of the low sintering temperature of the gold ink, conductive structures could be realized on COC. For improving the wettability of the gold ink on COC a low pressure oxygen plasma treatment was required. The adhesive strength of the gold ink could be improved by increasing the sintering temperature on COC. The plasma power has shown an influence on the adhesive strength, too. The resistance could be reduced by performing a photonic curing after the thermal sintering. The best results were reached with a plasma power of 40 W for 20 s and a sintering temperature of 130 ◦C with a subsequently performed photonic curing (Energy: 955 mJ/cm2). With the optimized process parameters, an electrochemical biosensor was successfully realized. The biosensor contained a working and counter electrode printed of gold. The reference electrode was printed of silver. Due to the poor adhesion of the printed silver layer, a gold layer was printed as the adhesion layer. Analysis of the effective surface revealed that the resulting structures are porous. Even though the porosity of the electrodes had no enhancing effect on DNA sensing in this work, other applications (e.g., glucose sensing [45]) may benefit from the relatively simple production process, which can be an attractive alternative to common fabrication methods like dealloying, template based synthesis [45] or chemical conversion of inkjet-printed Ag structures, which leads to only weakly cohesive porous gold [46]. The final DNA hybridization experiments demonstrate the principal applicability of inkjet-printed electrodes for the specific electrochemical detection of labelled DNA probes in a typical concentration and reaction buffer as used for nucleic acid amplification. Signals are comparable to those generated with commercial SPEs, but IPEs feature the advantage of a reduced pretreatment procedure. The printing of electrodes in a fluidic chamber as demonstrated should facilitate and cheapen the integration of electronics in microfluidic applications. Supplementary Materials: The following are available online at http://www.mdpi.com/1424-8220/20/5/1333/s1: Figure S1: Waveform of the Suntronic EMD5730; Figure S2: Waveform of the DryCure Au-J; Figure S3: Waveform of the XP PriElex® SU-8; Figure S4: Printing layouts for the optical and electrical characterization; Figure S5: Design of the 2D electrode array; Figure S6: Schemes and photography of the 2.5D array; Figure S7: Cartridge for hybridization experiments with inkjet-printed and screen-printed 2D arrays; Figure S8: Dependency of Aoxide on the number of cyclic voltammetry (CV) cycles in H2SO4; Figure S9: Number of immobilized capture probes plotted versus the effective electrode area Aoxide; Figure S10: Exemplary CV signal of functionalized 3-layer IPE and SPE; Figure S11: Hybridized signal probes on 2.5D arrays; Figure S12: SWV signals of the hybridized MB labelled signal probe; Table S1: Oligonucleotide sequences. Author Contributions: Conceptualization, M.T. and D.J.; methodology, M.T. and D.J.; validation, M.T. and D.J.; investigation, D.J. for inkjet-printing, M.T. and Z.B. for electrochemical DNA detection; writing—original draft preparation, M.T. and D.J.; writing—review and editing, N.B., T.M., F.v.S. and A.Z.; visualization, M.T. and D.J.; supervision, F.v.S.; project administration, K.G.; funding acquisition, F.v.S. and T.M. All authors have read and agreed to the published version of the manuscript. Funding: This research was funded by the German Federal Ministry for Economic Affairs and Energy (BMWi) through the Central Innovation Program for small and medium-sized enterprises (ZIM), grant numberPDF Image | Inkjet-Printing Nanoparticle Gold Silver Ink Cyclic Olefin
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