DSODARPA Silicon-based Ion Channel Sensor M. Goryll 1, S. Wilk 1, G. M. Laws 1, T. J. Thornton 1, S. M. Goodnick 1, M. Saraniti 2, J. Tang 3, R. S. Eisenberg.

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Presentation transcript:

DSODARPA Silicon-based Ion Channel Sensor M. Goryll 1, S. Wilk 1, G. M. Laws 1, T. J. Thornton 1, S. M. Goodnick 1, M. Saraniti 2, J. Tang 3, R. S. Eisenberg 3, 1 Arizona State University, Dept. of Electrical Engineering, Tempe, AZ 85287, 2 Illinois Institute of Technology, Dept. of Electrical and Computer Engineering, Chicago, IL Process Flow Project GoalsTechnical ApproachAchievements 150  m wide aperture in a silicon wafer, back side view AgCl Electrode Oxide SU-8 Resist Si Important building blocks of a fully integrated biosensor with on-chip sensing and signal processing Small Hole Etching 825 Resist, 1  m thickness AZ 4330 Resist, 2.6  m thickness Si Substrate 50  m 300  m SU-8 Resist Si 1 mm 250  m Si 150  m Si Thermally Grown Oxide, d = 500 nm Si 150  m Si Photoresist SU-8 Resist Si AgCl Hydrophobic Layer SU-8 Resist Si AgCl Bilayer Resist for Initial Hole Etching Thermal Oxidation Resist for Small Hole Etching Large Hole Etching SU-8 Resist (Epoxy) Surface Modification Layer AgCl Electrode AgCl Electrode, up to 1  m thickness SU-8 Resist Si Lipid Bilayer Attachment Experiment showing the opening of a single OmpF porin channel. The vertical lines through the red current trace are an artifact from stirring of the bath to facilitate the insertion of porin into the bilayer membrane. Plot showing the different levels of OmpF porin (Trimer). Level 1 is not shown. All the traces in the above plot are from the same OmpF porin bilayer experiment using the silicon wafer coated with PTFE (Teflon). Hole diameter = 150  m PTFE coated surface For the fabrication … coating the oxide surface with a Teflon film changes its properties from hydrophilic to hydrophobic (small to large contact angle) Plasma CVD provides an easy way of depositing several nm thick PTFE layers good agreement between model and experimental ellipsometric data allows a reliable thickness measurement dispersion curve indicates a high density PTFE polymer layer similar to bulk material “stackable” layers Bulk PTFE DuPont : n = 1.35 MIT : n = 1.38 PTFE layer on Si Index of refraction (n) Wavelength (nm) 900 Å layer 600 Å layer PTFE on Si: d = 598 Å ± 2 Å, n = ± 2E-3 ,  (degrees) Wavelength (nm) Model Fit (  in degrees) Model Fit (  in degrees) Exp  -E 75° (  in degrees) Exp  -E 75° (  in degrees) Blanket silver deposition by thermal evaporation on oxidized Si wafer Chloridization in 5% NaOCl minimal difference between the expected and measured Nernst potential variation with KCl concentration good potential stability of the microstructured electrodes Summary Silver Chloride Electrode the stability of the lipid bilayer is related to the contact angle between the bilayer and the supporting substrate water contact angle measure- ments can be used to determine the substrate’s surface energy  Bilayer Torus PTFE Surface Modification 3 Rush Medical College, Dept. of Molecular Biophysics and Physiology, Chicago, IL Plasma CVD is an interesting novel method to provide PTFE surface modification layers for lipid bilayer attachment to solid supports 0.5M (trans) and 0.6M (cis ) KCl Test solutions AgCl layer, chloridized in 5% NaOCl Potential difference (mV) Time (h) Simulation Measurement Potential difference (mV) KCl Molarity difference (M) AgCl Electrode Potential, Single substrate response is indistinguishable from channels in Teflon supported membranes, shows physiological behavior of OmpF reproducibility of measurements and voltage dependence indicates that switching is not an artifact but real channel activity switch to double-side polished 100 mm (4”) wafer with 380  m thickness allows the fabrication of multiple samples per run with identical geometry optimized backside alignment results in good centering of the hole front and backside have a smooth surface and the etching does not roughen the lower surface 150  m wide aperture in a silicon wafer, back side view (FESEM closeup) 150  m wide aperture in a silicon wafer, front side view measure single channels in an integrated device study the relation between the size of the lipid bilayer and the signal-to- noise ratio find optimal surface treatment for bilayer attachment find simulants that bind and transiently block conduction of ions through ompF work with DARPA and other groups MOLDICE groups to incorporate ion channels that show desired properties Future work under Phase IAccomplishments a silicon bilayer support chip has been constructed and successful Gigaseal formation has been demonstrated channel insertion succeeded first milestones have been achieved integration of the reversible electrodes demonstrated PTFE layers deposited by plasma CVD exhibit excellent properties Goal 1: Embed channels in an integrated device that maintains stable potential across them and allows recording of stable, artifact free current through them. Goal 2: Find simulants that bind and transiently block conduction of ions through OmpF.* * we shall work with DARPA and other groups within the MOLDICE network to incorporate ion channels that show desired properties silicon substrates are used layers are structured by conventional optical lithography the aperture that supports the bilayers is constructed using deep silicon dry etching relation between the size of the lipid bilayer and its stability and the signal-to-noise ratio of the ion channel response ultimate limit for the size scaling of the sensor optimal surface treatment for lipid bilayer attachment stability of the integrated reversible Ag/AgCl electrodes manufacturability of the sensor usability issues (reusability, cleaning) Challenges we are facing impedance analysis of bilayers current-voltage measurements of bilayers and porin channels studying the influence of surface modification layers on bilayer Gigaseal formation Experiments involve … maintain stable potential (± 1 mV for 1 hour) across a single channel of OmpF porin recording of stable, artifact - free current voltage curves (± 100 pA for 1 hour) from a single channel of OmpF porin using external electrodes recording stable current voltage curves using integrated Ag/AgCl electrodes Milestones measure sealing resistance on samples with different geometries and surface properties measure Nernst potential of Ag/AgCl electrodes measure DC potential across porin measure current through porin Demonstration of Results