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Presented at N2L meeting, Lund, October 2007. Whole cell biochips – Issues in Nano Bio Interfacing . Yosi Shacham-Diamand. The Bernard L. Schwartz chair for nano Scale information Technologies The Dept. of Physical Electronics , School of EE , Faculty of Engineering

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slide1

Presented at N2L meeting, Lund, October 2007

Whole cell biochips – Issues in Nano Bio Interfacing

Yosi Shacham-Diamand

The Bernard L. Schwartz chair for nano Scale information Technologies

The Dept. of Physical Electronics ,

School of EE , Faculty of Engineering

Tel-Aviv University, Israel

&

The Dept. of Applied Chemistry

Waseda University, Tokyo, Japan

biological recognition hierarchy
Biological Recognition Hierarchy

Antibody

Specificity

Enzyme

Complexity

Hierarchy

DNA

Whole cell

Physiological effect

Tissue

why integrating live cells
Why integrating live cells ?
  • Multi-cells:
  • Functional response
  • Emulating “real” life behavior
  • Emulating complex systems characteristics
  • Study cell behavior
  • Single cells:
  • All the above + cell sorting
slide4

“Canary in a cage” concept

Photograph from the "Welsh Coal Mines" Collection from the National Museum of Wales

whole cell bio chip
Whole cell Bio-Chip
  • Prokaryotic – Bacteria,
    • Sensors for acute toxicity in water,
    • Detecting toxicity of drugs, cosmetics etc.
  • Eukaryotic –
    • Mammalian cells - cancer therapy, stem cells characteristics
interfacing cell biology mems
Interfacing cell biology & MEMS
  • Optical
    • Luminescence – photo luminescence, bio-luminescence
  • Electrical
    • Electrochemical – active or passive electrodes
    • Impedance spectroscopy
  • Mechanical
    • Resonators, deflection sensors
why electrochemistry
Why Electrochemistry?
  • Simple.
  • Sensitive.
  • Monitoring in turbid solutions.
  • Simultaneous measurements of several samples.
  • Electrical output- convenient to handle and analyze.
  • Easily scaled down.
genetically engineered bacteria

Substrate

Bacteria

Promoter

Reporter

Plasmid

Toxicant

Genetically Engineered Bacteria

b-gal

b-gal

b-gal

genetically engineered bacteria10

PAPG

Genetically Engineered Bacteria

Bacteria

Enzyme Sabstrate

Product

PAP+

b-gal

genetically engineered bacteria11

PAPG

PAP

PAP

PAP

PAP

PAP

PAP

Genetically Engineered Bacteria

Bacteria

Enzyme Sabstrate

Product

+

b-gal

portable biochip system
Portable BioChip System

Array of nano liter volume electrochemical cells

Multiplexer

Pocket PC

Potentiostat

a single chamber

(Magnified)

slide19
Goal

Evaluation of cancer cells response to different drugs.

Bio-chips for differential therapy

introduction
Introduction

Current Therapeutic Strategies:

  • Surgery
  • Chemotherapy
  • Irradiation

Differentiation Therapy:

  • Cancerous cells are being induced to behave like normal cells
  • It restrains their growth
  • Differentiation agents tend to have less toxicity than conventional cancer treatments
how can we evaluate the efficiency of the drug
How can we evaluate the efficiency of the drug?

According to the enzymatic activity level of the treated cancer cells.

  • Normal enzymatic activity denotes that the cells become 'healthy‘,
  • Lack of enzymatic activity denotes ineffectual drug treatment for the particular cancer tumor and for the particular patient.
experimental
Experimental
  • Human colon cancer cells were treated with different differentiation therapy drug agents.
  • The cells were placed in each one of the electrochemical-cells in the array, while each chamber was treated with different drug type.
  • p-APP substrate was added.
  • The generated current signal was measured.
  • Cells number was counted under the microscope.
slide25

Correlation between HT-29 colon cancer cell number and the induced alkaline phosphatase enzymatic activity (DI/Dt). (amperometric signal at 220mV).

photo luminescent bio chip
Photo luminescent bio-chip

Emission (green)

Photodiode

Excitation chip (Blue)

Cell’s container

Bio chip

bio luminescent sensor
Bio-luminescent sensor

Emission (green)

Photodiode

Cell’s container

Bio chip

slide29

Reporting element

gene promoter

luxCDABE

Sensing element

Engineering live cells for the detection of toxicants

The fusion of two genetic elements:

  • Sensing element: A promoter of a geneinvolved in the response to the desired target
  • Reporting element:Fluorescence or bioluminescence genes

The final construct emits a dose-dependent signal in response to the presence of the target chemicals

Light

slide30

Bioluminescent Prokaryote cell-based biochip

  • Comprised of:
  • (a) biochip sensor for optical/electrochemical measurement.
  • (b) microfluidic elements for delivery of samples and media.
  • A nano-patterning technique for spotting bacteria onto a platform is being developed.
  • The biochip functions in a “plug-&-play” mode of action to facilitate insertion into the microfluidic platform.
slide31

System outline

Prokaryote cell biochip layout

Schematic view of the four photo-diodes array

slide32

New setup – with 4 PV detectors/ mechanical scan

Biochip platform (on the left) and its 3D model (on the right).

slide34

Sigmoid

Cross-correlation:

10 ppm = 0.631

5 ppm = 0.647

more complicated systems bio mems lab on chip
More complicated systemsBio-MEMS Lab-on-Chip
  • Using MEMS technology integrating low-light emitting whole-cell sensors, and VLSI devices.
  • Micromechanical shutters for luminescent bio-chips – modulates the light

Optical Sensor

Modulation

Shutters

Luminescence

Whole cell Biochip

more complicated systems bio mems lab on chip38
More complicated systemsBio-MEMS Lab-on-Chip
  • Using MEMS technology integrating low-light emitting whole-cell sensors, and VLSI devices.
  • Micromechanical shutters for luminescent bio-chips – modulates the light

Optical Sensor

Modulation

Shutters

Luminescence

Whole cell Biochip

more complicated systems bio mems lab on chip39
More complicated systemsBio-MEMS Lab-on-Chip
  • Using MEMS technology integrating low-light emitting whole-cell sensors, and VLSI devices.
  • Micromechanical shutters for luminescent bio-chips – modulates the light

Optical Sensor

Modulation

Shutters

Luminescence

Whole cell Biochip

more complicated systems bio mems lab on chip40
More complicated systemsBio-MEMS Lab-on-Chip
  • Using MEMS technology integrating low-light emitting whole-cell sensors, and VLSI devices.
  • Micromechanical shutters for luminescent bio-chips – modulates the light

Optical Sensor

Modulation

Shutters

Luminescence

Whole cell Biochip

slide41

Integrated heterodyne detection with Whole cell biochips

Heterodyne detection

Output

Optical Sensor

Shutters

Modulator

~1kHz

Luminescence

Whole cell Biochip

  • Converts low frequency biological signal to high frequency signal,
  • Reduces 1/f noise  improves the S/N ratio.
fabrication results

500 μm

200 μm

Fabrication Results

Shutters, Springs, Comb-drives Backbone, Shutters, Shutter-Windows

slide43

MEMS Fabrication

Array of resonators as band-pass filters Array of comb-drive actuators

slide44

Fabrication Results

Released actuators Cross-section of electrically isolated device

fabrication results backside characterization
Fabrication Results: Backside Characterization

Goal: Characterize Deep Silicon backside etch of the shutter windows using the Bosch Process using windows with varying gaps

Gap: 65 μm

Gap: 60 μm

Gap: 70 μm

Gap: 50 μm

Gap: 45 μm

Gap: 55 μm

cross section sketch showing the components of the experimental set up
Cross-section sketch showing the components of the experimental set-up.

Light emitted from the bio chip

frequency response of the device
Frequency response of the device.

In air In vacuum

Noel Elman, PhD thesis , TAU 2006

integrated heterodyne mems
Integrated Heterodyne MEMS

Response vs. Concentration

Response vs. time

key issues
Key issues
  • Cell storage on the chip
  • Cell revival
  • Signal level – very low, the microbes emit ~ 0.1 – 10 photons/ sec.
  • Operation under flowing liquid
  • Detection in air – extracting onto water
  • Producing arrays
acknowledgements
Acknowledgements

Thanks to all my students, especially to Dr. Rachela Popovtzer (Graduated 2006) , Dr. Noel Elman (Graduated 2006), Dr. Ronen Almog (Post Doc), Arthur Rabner (2009), Hadar Ben-Yoav (2009), Sefi Wornick (2009), Amit Ron (2009), Amit Livneh (2007), Hila Einati (2009) and Hila Dagan (2008)

Special thanks to Prof. Shimshon Belkin from the Hebrew University of Jerusalem (HUJI)

Prof. Judith Rishpon and Prof. Eliora Ron from Tel Aviv University (TAU)

Dr. Slava Krylov (TAU) and Dr. Marek Sternhaim for their help with the MEMS modeling

thanks
Thanks

Zichron -Yaakov