Nano and Microtechnologies of hybrid bioelectronic systems Nano and Microtechnologies of hybrid...

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Transcript of Nano and Microtechnologies of hybrid bioelectronic systems Nano and Microtechnologies of hybrid...

  • Slide 1
  • Nano and Microtechnologies of hybrid bioelectronic systems Nano and Microtechnologies of hybrid bioelectronic systems (Lecture 2- nano-topography) Dr. Yael Hanein
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  • Nano and Microtechnologies of hybrid bioelectronic systems Cell Patterning Approaches Direct protein lithography Micro-contact printing/micro fluidics Proteins SAMs Dry lithography Patterned polymers Temperature sensitive polymers Nano-topography Ordered nano-patterning Disordered nano-patterning Wells
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  • Nano and Microtechnologies of hybrid bioelectronic systems Approaches (V) : Nanotopography Ancient methods Micro-methods Silicon pillars Silicon grass Nano-methods Carbon nanotubes
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  • Nano and Microtechnologies of hybrid bioelectronic systems Approaches (V) : Nanotopography Thermally grown SiO2 Resist Exposure, development RIE, CHF3: oxide etch Photoresist removal RIE: Cl2, BCl3 Si etch HF: Oxide removal
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  • Nano and Microtechnologies of hybrid bioelectronic systems Approaches (V) : Nanotopography http://www.hgc.cornell.edu/neupostr/lrie.htm
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  • Nano and Microtechnologies of hybrid bioelectronic systems Approaches (V) : Nanotopography http://www.wadsworth.org/divisions/nervous/nanobio/DG06.htm
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  • Nano and Microtechnologies of hybrid bioelectronic systems Approaches (V) : Nanotopography Craighead RIE: Cl 2,CF 4,O 2 Photoresist Wet etching: HF, nitric acid, H 2 O Resist removal, Cleaning
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  • Nano and Microtechnologies of hybrid bioelectronic systems Approaches (V) : Nanotopography LRM55 Astroglial cells prefer smooth surfaces Cortical astrocytes Preferred rough surface
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  • Nano and Microtechnologies of hybrid bioelectronic systems Culture of neural cells on silicon wafers with nano-scale surface topograph Y.W. Fan et all, Culture of neural cells on silicon wafers with nano- scale surface topograph : Si surfaces with variable roughness (without surface treatment) -Morphology of adherent cells remarkably differs on differently rough surfaces Larger contact area? doesnt explain the decline in cell adhesion after a certain Ra value! (Can you really change Ra without changing other parameters?!)
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  • Nano and Microtechnologies of hybrid bioelectronic systems Cells and nanotopography Cells respond to surface topography The mechanisms involving cell adhesion and migration on surfaces is poorly understood Extremely important in the field of tissue engineering and biomaterials Important in lab-on a chip/micro bio-sensors
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  • Nano and Microtechnologies of hybrid bioelectronic systems Cells React to Nanoscale Order and Symmetry in Their Surroundings A. S. G. Curtis*, N. Gadegaard, M. J. Dalby, M. O. Riehle, C. D. W. Wilkinson, and G. Aitchison
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  • Nano and Microtechnologies of hybrid bioelectronic systems Methods Arrays of nano-pits were prepared in a three-step process: Electron Beam Lithography Nickel die fabrication Hot embossing into polymers
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  • Nano and Microtechnologies of hybrid bioelectronic systems Electron Beam Lithography Positive resist ZEP 520A coating on silicon EBL of pits, with diameter of 35, 75, 120 nm Development
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  • Nano and Microtechnologies of hybrid bioelectronic systems Nickel die fabrication 100 nm thick resist with nanopits Sputter coating of a 50nm Ni-V laye Electroplating of Ni to a thickness of 300 m Nickel Die
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  • Nano and Microtechnologies of hybrid bioelectronic systems Hot embossing into polymers Polymeric replicas were made by embossing the nickel die in a heated polymethylmethacrylate (PMMA) or polycaprolactone (PCL) sheets
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  • Nano and Microtechnologies of hybrid bioelectronic systems Cell Cultures Primary human fibroblasts (connective tissue cells)/ rat epithenon cells were seeded on patterned PCL or PMMA 1.Short term experiments: measurements taken at intervals from 2-24 hr 2.Long term experiments: cells cultured for up to 71 days counting n o of adherent cells and measuring their orientation
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  • Nano and Microtechnologies of hybrid bioelectronic systems Adhesion on spaced nanopatterened areas is much lower than on planar areas, but on the smallest closest spaced pits is the same as on the planar area! Rat epitenon cells grown on PCL surfaces for 24 h Human fibroblast cells grown on PCL
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  • Nano and Microtechnologies of hybrid bioelectronic systems Many cells possess surface nanometric features Filopodia and microspikes may be the organelle whose major function is to explore nanofeatures around the cell It is interesting to note that the filopodia follows the nanopattern, and seems to be directed by it
  • Slide 19
  • Nano and Microtechnologies of hybrid bioelectronic systems Reaction of cells to different symmetries Cathrine C. Berry et all, The influence of microscale topography on fibroblast attachment and motility: fibroblasts were grown on arrays of pits, 7, 15 and 25 diameter, 20 and 40 m spacing 1.Cells prefer entering the larger diameter pits, meaning they might be sensitive to differences in radius of curvature 2.The smallest pits allow the highest proliferation rate and the highest migration rate of a single cell
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  • Nano and Microtechnologies of hybrid bioelectronic systems On orthogonal patterns :cells show preference of 90 separated orientations On hexagonal patterns: cells show preference of 120 separated orientations Orientation is nonrandom Cells can distinguish between symmetries???
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  • Nano and Microtechnologies of hybrid bioelectronic systems Fredrick Johansson et al, Axonal outgrowth on nano-imprinted patterns Investigated guidance of axons on patterns of parallel grooves of PMMA, with depths of 300nm, widths of 100-400 nm and distance between grooves 100-1600 nm. -axons display contact guidance on all patterns -preferred to grow on edges and elevations in the patterns rather than in grooves- this may be due to edge effects, as concentration of charges
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  • Nano and Microtechnologies of hybrid bioelectronic systems What makes cells adhere to surfaces ? How cells sense ORDER and SYMMETRY of surfaces? Why do differences in diameters and spacing of micro and nano features have such dramatic effect on cell adhesion?
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  • Nano and Microtechnologies of hybrid bioelectronic systems Two possible explanations The effect is caused by the nonliving surfaces alone Nanofeatures are known to affect orientations in nonliving systems It is unknown whether nanofeatures affect protein adsorption on the nanoscale, (exposure to protein rich culture media- showed no difference) The effect is caused by interaction of cellular processes and interfacial forces
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  • Nano and Microtechnologies of hybrid bioelectronic systems Ordered conducting grooves Rough conducting substrate Random nano-topography insulating substrate Ordered insulating grooves Perturbed ordered insulating grooves Nano topography Types of nano-topography
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  • Nano and Microtechnologies of hybrid bioelectronic systems Ra Symmetry
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  • Nano and Microtechnologies of hybrid bioelectronic systems Carbon nanotube based neuro- chips for engineering and recording of cultured neural networks
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  • Nano and Microtechnologies of hybrid bioelectronic systems Recording from cultured neural networks Ben-Jacob, TAUFromherz, MPIBauman, URosGross, UNT
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  • Nano and Microtechnologies of hybrid bioelectronic systems Multi electrode arrays E. Ben-Jacob Large electrically active networks, Long term (weeks), Relevant biological activity BUT Large electrodes, Poor sealing, Average (many neurons) signal, Poor electrode-cell coupling, Random networks
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  • Nano and Microtechnologies of hybrid bioelectronic systems Multi electrode arrays
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  • Nano and Microtechnologies of hybrid bioelectronic systems Outline How can we make better/new MEA How do we manipulate cells on substrates? Properties of our new MEAs
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  • Nano and Microtechnologies of hybrid bioelectronic systems How can we make better/new MEA? Signal fidelity ~ Noise CeCe ReRe C sh R spread R met R seal C hd Soma CeCe ReRe C sh R spread R met R seal C hd Kovacs, 1994
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  • Nano and Microtechnologies of hybrid bioelectronic systems Cell-substrate interactions Wong et al. Surface chemistry 2004
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  • Nano and Microtechnologies of hybrid bioelectronic systems Nano-topography Hu et al. Mattson et al. J. Mol. Neurosci 2000 Craighead, Cornell
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  • Nano and Microtechnologies of hybrid bioelectronic systems Electronic properties (CNTs) armchair zigzag
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  • Nano and Microtechnologies of hybrid bioelectronic systems Carbon nanotubes Biocompatible Super capacitors Compatibility with micro fabrication CNT electrodes Self-cell-organization Network engineering Excellent recording Carbon nanotube multi-electrode arrays
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  • Nano and Microtechnologies o