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Detection of Pathogens Using Electrochemical DNA Sensors for Resource-Limited Settings
Sarah Ghanbari, Nicholas Giustini,
Cameron Mar, Pankti Doshi, Unyoung Kim
Santa Clara University
October 15th, 2011
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Overview
Problem Statement
Current Technological Solutions
Key Components of Design– High throughput concentrator– Lysis Chamber– DNA Sensor Chamber
Concentrator– Fabrication– Analysis
Sensor Chamber– Analysis
Summary of Work
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Problem Statement
Approximately one in eight people lack access to safe water.1
The water and sanitation crisis claims more lives through disease than any war claims through guns (with more than 3.5 millions deaths each year).2
Diarrhea is the second leading cause of death in children under five. It kills more young children than AIDS, malaria, and measles combined.3
1. Special Focus on Sanitation. UNICEF, WHO. 2008. 2. 2006 United Nations Human Development Report. 3. Diarrhea: Why children are still dying and what can be done. UNICEF, WHO 2009.
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Problem Statement
Populations without access to safe drinking water
No Data1% - 25%
26% - 50% 51%-75% 76% - 100%
The World’s Water The Biennial Report on Freshwater Resources (Gleick 1998)
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Research Question
Can we make a device that is:– Small and portable– Reduces reagent and power consumption– More accurate– Provides faster diagnosis– User friendly
Yes, by utilizing a microfluidic platform
Our method is to create a microfluidic platform combining an inertial concentrator and electrochemical DNA sensor.
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Problems with Current Solutions
Time consuming
Expensive
Tests only indicate possibility of contamination
1.Potatest water test kit by Wagtech WTD2. Sengupta, Shramik , et. al, Microfulidic Diagnostic Systems.
1
Traditional Tests Developing Microfluidic Tests2
Complicated fabrication and architecture
Expensive and time consuming preparatory procedures
Requires non-portable equipment for full functionality
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Key Components of Design
High-throughput concentrator
Cell lysis chamber
Electrochemical DNA sensor
Schematics of High-throughput Concentrator
Schematics of Electrochemical DNA Sensor
Inlet
Outlet
*Di Carlo, Dino D., et al, PNAS Vol. 104, pp. 18892-18897 (2007)
Rf = 2ra2/Dh3
High current
Lowcurrent
Lysis Chamber
Rf : Inertial Force Ratior : Radius of Curvaturea : Particle SizeDh: Hydraulic Diameter
*
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Remove Uncured Monomer
Fabrication:
Photolithography
Etching
Resist Removal
Wafer Bonding
Coat Substrate
Fill Chamber with Monomer Mixture
Align Photomask
Expose with UV
*Hutchison, J. Brian, et. al., Lab on a Chip 4.6 (2004): 658-62.
Traditional Methods Contact Liquid Polymer Process(CLiPP)*
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Concentrator Results
Flow Rate: 0.1 mL/min
Flow Rate: 1.6 mL/min
Flow Rate:1.1 mL/min
Flow Rate:0.2 mL/min
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Chemistry of the DNA Sensor
Thiol attachment to goldMethylene Blue
Methylene Blue
Self Hybridization Region
SensorProbe
SensorProbe
TargetSequence
Au Working
Electrodes
Pt Counter &Reference
Thiol attachment to gold
High current
Lowcurrent
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High current
Lowcurrent
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Summary
Key components– Concentrator: achieved focusing for 10 μm particles at a flow rate
of 1.1 mL/min to concentrate the pathogens in a water sample– DNA Sensor Chamber: confirmed the specificity of DNA sensor
strands toward an identified target (E. coli) at a concentration of 500 nM
Ongoing and Future Work– Improve concentrating efficiency for 0.5-2 µm particles and
asymmetrical particles – Integrate concentrator, lysis chamber, and sensor chamber into a
monolithic chip
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Acknowledgements
Dr. Ashley Kim
Dr. Teresa Ruscetti
Dr. Steven Suljak
Mr. Daryn Baker
Dr. Cary Yang
Dr. Dan Strickland
Dr. Hohyun Lee
Stanford Nanofabrication Facilities
Roelandts Fellows
School of Engineering
Biomedical Engineering Society
OAI
