Whole cell biochips

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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 Tel-Aviv University, Israel
The Dept. of Applied
Chemistry Waseda University, Tokyo, Japan
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Biological Recognition Hierarchy
Antibody
Specificity
Enzyme
Complexity Hierarchy
DNA
Whole cell
Physiological effect
Tissue
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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

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Canary in a cage concept
Photograph from the "Welsh Coal Mines" Collection
from the National Museum of Wales
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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

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Interfacing cell biology MEMS

• Optical
• Luminescence photo luminescence,
  bio-luminescence
• Electrical
• Electrochemical active or passive electrodes
• Impedance spectroscopy
• Mechanical
• Resonators, deflection sensors

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Electrochemical whole cell bio-chips
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Why Electrochemistry?

• Simple.
• Sensitive.
• Monitoring in turbid solutions.
• Simultaneous measurements of several samples.
• Electrical output- convenient to handle and
  analyze.
• Easily scaled down.

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Genetically Engineered Bacteria
b-gal
b-gal
b-gal
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Genetically Engineered Bacteria
Bacteria
Enzyme Sabstrate
Product
PAP
b-gal
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Genetically Engineered Bacteria
Bacteria
Enzyme Sabstrate
Product

b-gal
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Chip Process
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Chip Process
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Chip Process
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Plastic Platform
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Portable BioChip System
Array of nano liter volume electrochemical cells
Multiplexer
Pocket PC
Potentiostat
a single chamber (Magnified)
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Physiological Response To Phenol On-chip
Control
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Personalized MedicineHigh-throughput Detection
Of Human Cancer Cells
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Goal
Evaluation of cancer cells response to different
drugs. Bio-chips for differential therapy
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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

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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.

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Experimental

1. Human colon cancer cells were treated with
   different differentiation therapy drug agents.
2. The cells were placed in each one of the
   electrochemical-cells in the array, while each
   chamber was treated with different drug type.
3. p-APP substrate was added.
4. The generated current signal was measured.
5. Cells number was counted under the microscope.

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Results
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Enzymatic activities vs. cancer cell number
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Correlation between HT-29 colon cancer cell
number and the induced alkaline phosphatase
enzymatic activity (DI/Dt). (amperometric signal
at 220mV).
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Optical whole cell bio-chips
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Photo luminescent bio-chip
Emission (green)
Photodiode
Excitation chip (Blue)
Cells container
Bio chip
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Bio-luminescent sensor
Emission (green)
Photodiode
Cells container
Bio chip
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Engineering live cells for the detection of
toxicants
The fusion of two genetic elements

• Sensing element A promoter of a gene involved
  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
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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.

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System outline
Prokaryote cell biochip layout
Schematic view of the four photo-diodes array
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New setup with 4 PV detectors/ mechanical scan
Biochip platform (on the left) and its 3D model
(on the right).
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(No Transcript)
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Sigmoid
Cross-correlation 10 ppm 0.631 5 ppm 0.647
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Verhulst
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Low level signal detection
60uL , NA 16ppm, IT100msec
500uL, NA 3ppm, PD Area 1mm2
Photo multiplier Photo diode
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System optimization using the ASAP sofware
ra0.5 mm
Total bacteria 3.5x105 CFU
Optimal detector radius
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Integrated heterodyne detection with Whole cell
biochips
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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
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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
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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
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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
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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.

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Fabrication Results

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

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MEMS Fabrication

Array of resonators as band-pass filters
Array of comb-drive actuators
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Fabrication Results
Released actuators
Cross-section of electrically isolated device
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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
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Cross-section sketch showing the components of
the experimental set-up.
Light emitted from the bio chip
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Frequency response of the device.
In air
In vacuum
Noel Elman, PhD thesis , TAU 2006
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Response to IPTG (isopropyl-beta-D-thiogalactopyra
noside) (0.1 mM)
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Integrated Heterodyne MEMS
Response vs. Concentration
Response vs. time
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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

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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
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Acknowledgement
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Thanks
Zichron -Yaakov