微流控芯片之一
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– Portable devices for pathogen detection – High-throughput biology
• • • • • • •
Chemical synthesis Cell Immunoassay analysis Metabolite analysis Drug delivery systems Single molecular Enzyme analysis
Lab on a Chip
Or: The Incredible Shrinking Science Experiment! Or: APOIBE!
Credits: Mathies Lab, UC-Berkeley Quake Lab, Stanford Agilent, Inc.
二十一世纪科学发展趋势
Thermal Cycling Characteristics
Temperature (°C)
Time (seconds)
Time (seconds)
> 5X faster than conventional thermal cyclers
Application: Tuberculosis (TB) detection
D. W. Inglis, J. A. Davis, T. J. Zieziulewicz, D. A. Lawrence, R. H. Austin, and J. C. Sturm, Determining blood cell size using microfluidic hydrodynamics, J. Immunol. Meth., vol. 329, pp. 151-156, 2008.
Yeast Blood Cells cryptosporidium
Microfluidic or Lab-on-a-chip Devices can be used to separate different cell types
Various blood cell separations have been demonstrated:
But no work yet with microorganisms!
The Not-so-Distant Future
2Байду номын сангаас08
PDA
2308??
Paramount
384根通道扫描检测
Emrich, C.A., et al.,Analytical Chemistry, 2002. 74(19): p. 5076-5083.
10根通道CCD同时检测
Dang, F.Q., et al., Analytical Chemistry, 2005. 77(7): p. 2140-2146.
Credit: Dr. Karen Cheung, UBC ECE
Why use microfluidics?
Credit: Dr. Karen Cheung, UBC ECE
1 molecule in 1 mL = 1.6 x 10-18 M 1 molecule in 1 nL = 1.6 x 10-15 M 1 molecule in 1 pL = 1.6 x 10-12 M
Disease caused by Mycobacterium tuberculosis 9 million new cases of active TB every year worldwide
2 million TB deaths every year
450,000 new cases of multi-drug resistant TB / yr
目前最高通量微流控芯片平台
Aborn, J.H., et al., Lab on a Chip, 2005. 5(6): p. 669-674
门控进样多通道检测
Xu, X.M., et al., Analytical Chemistry, 2002. 74(21): p. 5517-5522
高通量均相免疫分析
Why use microfluidics?
• Sample savings – nL of enzyme, not mL • Faster analyses – can heat, cool small volumes quickly • Integration – combine lots of steps onto a single device • Novel physics – diffusion, surface tension, and surface effects dominate
Bromberg, A. and R.A. Mathies, Electrophoresis, 2004. 25(12): p. 1895-1900.
Lab-on-a-chip Devices for Cell Separation and Identification
Microbes, cells and small organisms come in a wide array of shapes and sizes
Bacterial Pathogens
Food poisoning
Wound infections Lung Disease
Dr. Sharon Peacock, University of Oxford
Biowarfare
Food poisoning: Pathogenic E. coli: >1 in 10,000 in Canada Salmonella spp.: as high as 1 in 50 eggs Wound infections: Antibiotic-resistant S. aureus: as high as 50%
10% of TB cases in South Africa are XDR TB strains, resistant to first- and second-line antibiotics
Doctors Without Borders:
Current TB detection methods
Develop bacterial pathogen detectors that are:
Rapid (< 1 hour)
General (many different types of pathogens)
Sensitive (detect a single cell)
Genetically Specific
– This can actually lead to faster reactions!
Gel electrophoresis
1-3ｈ
Capillary electrophoresis
sample separation capillary detection
buffer
High voltage
buffer
• • • • Miniaturization Automation Integration Portable
Lab on a Chip?
Microfluidics
Microfluidics: The use of microfabrication techniques from the IC industry to fabricate channels, chambers, reactors, and active components on the size scale of the width of a human hair or smaller
Lung Disease: Pseudomonas spp. Mycobacterium tuberculosis
Biowarfare: Anthrax, plague, smallpox
Methods for Genetic Detection of Pathogens
OLD WAY:
NEW WAY:
Courtesy of Dr. Boris Stoeber
Genetic analysis microsystems
96-channel radial capillary array electrophoresis microplate
Anal Chem. 1999 ;71(23):5354-61
• J. A. Davis, D. W. Inglis, K. M. Morton, D. A. Lawrence,
L. R. Huang, S. Y. Chou, J. C. Sturm, and R. H. Austin, Deterministic hydrodynamics: taking blood apart, Proc. Nat. Acad. Sci. (USA), vol. 103, pp. 14779-14784, 2006.
Field-portable
PCR-CE Microsystem Fabrication
Portable System for Infectious Disease Detection
System measures 8 x 10 x 12 inches
•
• • • •
Complete fluorescence detection including 488-nm solid-state laser, PMT, optics, and detection electronics Heater and RTD electronics 4 high-voltage CE power supplies PDMS microvalve hardware Simple PC interface
10-30 min
Microchip electrophoresis
1-3 min Multi-channel microchip electrophoresis
1-3 min/ 12 samples
Some interesting LOC applications • Genetic analysis
1. Fixation and staining with acid-fast stains (Ziehl-Neelsen or auramine stains) 2. Optical or fluorescence microscope examination ( 10,000 organisms/mL)
•D. W. Inglis, K. J. Morton, J. A. Davis, T. J. Zieziulewicz, D. A. Lawrence, R. H. Austin, and J. C. Sturm, Microfluidic device for label-free measurement of platelet activation, Submitted to Lab on a Chip, Jan 08.
3. Culture in biosafety level 3 containment facility
In select cases, molecular detection is used to verify diagnosis "That TB destroys millions of lives around the world every year shows that the current approach is just not working…The tools we have to treat and diagnose TB are woefully inadequate and outdated, and we're not seeing the necessary urgency to tackle the disease." Dr. Tido von Schoen-Angerer, Director, DWB’s Campaign for the Access to Essential Medicines
• • • • • • •
Chemical synthesis Cell Immunoassay analysis Metabolite analysis Drug delivery systems Single molecular Enzyme analysis
Lab on a Chip
Or: The Incredible Shrinking Science Experiment! Or: APOIBE!
Credits: Mathies Lab, UC-Berkeley Quake Lab, Stanford Agilent, Inc.
二十一世纪科学发展趋势
Thermal Cycling Characteristics
Temperature (°C)
Time (seconds)
Time (seconds)
> 5X faster than conventional thermal cyclers
Application: Tuberculosis (TB) detection
D. W. Inglis, J. A. Davis, T. J. Zieziulewicz, D. A. Lawrence, R. H. Austin, and J. C. Sturm, Determining blood cell size using microfluidic hydrodynamics, J. Immunol. Meth., vol. 329, pp. 151-156, 2008.
Yeast Blood Cells cryptosporidium
Microfluidic or Lab-on-a-chip Devices can be used to separate different cell types
Various blood cell separations have been demonstrated:
But no work yet with microorganisms!
The Not-so-Distant Future
2Байду номын сангаас08
PDA
2308??
Paramount
384根通道扫描检测
Emrich, C.A., et al.,Analytical Chemistry, 2002. 74(19): p. 5076-5083.
10根通道CCD同时检测
Dang, F.Q., et al., Analytical Chemistry, 2005. 77(7): p. 2140-2146.
Credit: Dr. Karen Cheung, UBC ECE
Why use microfluidics?
Credit: Dr. Karen Cheung, UBC ECE
1 molecule in 1 mL = 1.6 x 10-18 M 1 molecule in 1 nL = 1.6 x 10-15 M 1 molecule in 1 pL = 1.6 x 10-12 M
Disease caused by Mycobacterium tuberculosis 9 million new cases of active TB every year worldwide
2 million TB deaths every year
450,000 new cases of multi-drug resistant TB / yr
目前最高通量微流控芯片平台
Aborn, J.H., et al., Lab on a Chip, 2005. 5(6): p. 669-674
门控进样多通道检测
Xu, X.M., et al., Analytical Chemistry, 2002. 74(21): p. 5517-5522
高通量均相免疫分析
Why use microfluidics?
• Sample savings – nL of enzyme, not mL • Faster analyses – can heat, cool small volumes quickly • Integration – combine lots of steps onto a single device • Novel physics – diffusion, surface tension, and surface effects dominate
Bromberg, A. and R.A. Mathies, Electrophoresis, 2004. 25(12): p. 1895-1900.
Lab-on-a-chip Devices for Cell Separation and Identification
Microbes, cells and small organisms come in a wide array of shapes and sizes
Bacterial Pathogens
Food poisoning
Wound infections Lung Disease
Dr. Sharon Peacock, University of Oxford
Biowarfare
Food poisoning: Pathogenic E. coli: >1 in 10,000 in Canada Salmonella spp.: as high as 1 in 50 eggs Wound infections: Antibiotic-resistant S. aureus: as high as 50%
10% of TB cases in South Africa are XDR TB strains, resistant to first- and second-line antibiotics
Doctors Without Borders:
Current TB detection methods
Develop bacterial pathogen detectors that are:
Rapid (< 1 hour)
General (many different types of pathogens)
Sensitive (detect a single cell)
Genetically Specific
– This can actually lead to faster reactions!
Gel electrophoresis
1-3ｈ
Capillary electrophoresis
sample separation capillary detection
buffer
High voltage
buffer
• • • • Miniaturization Automation Integration Portable
Lab on a Chip?
Microfluidics
Microfluidics: The use of microfabrication techniques from the IC industry to fabricate channels, chambers, reactors, and active components on the size scale of the width of a human hair or smaller
Lung Disease: Pseudomonas spp. Mycobacterium tuberculosis
Biowarfare: Anthrax, plague, smallpox
Methods for Genetic Detection of Pathogens
OLD WAY:
NEW WAY:
Courtesy of Dr. Boris Stoeber
Genetic analysis microsystems
96-channel radial capillary array electrophoresis microplate
Anal Chem. 1999 ;71(23):5354-61
• J. A. Davis, D. W. Inglis, K. M. Morton, D. A. Lawrence,
L. R. Huang, S. Y. Chou, J. C. Sturm, and R. H. Austin, Deterministic hydrodynamics: taking blood apart, Proc. Nat. Acad. Sci. (USA), vol. 103, pp. 14779-14784, 2006.
Field-portable
PCR-CE Microsystem Fabrication
Portable System for Infectious Disease Detection
System measures 8 x 10 x 12 inches
•
• • • •
Complete fluorescence detection including 488-nm solid-state laser, PMT, optics, and detection electronics Heater and RTD electronics 4 high-voltage CE power supplies PDMS microvalve hardware Simple PC interface
10-30 min
Microchip electrophoresis
1-3 min Multi-channel microchip electrophoresis
1-3 min/ 12 samples
Some interesting LOC applications • Genetic analysis
1. Fixation and staining with acid-fast stains (Ziehl-Neelsen or auramine stains) 2. Optical or fluorescence microscope examination ( 10,000 organisms/mL)
•D. W. Inglis, K. J. Morton, J. A. Davis, T. J. Zieziulewicz, D. A. Lawrence, R. H. Austin, and J. C. Sturm, Microfluidic device for label-free measurement of platelet activation, Submitted to Lab on a Chip, Jan 08.
3. Culture in biosafety level 3 containment facility
In select cases, molecular detection is used to verify diagnosis "That TB destroys millions of lives around the world every year shows that the current approach is just not working…The tools we have to treat and diagnose TB are woefully inadequate and outdated, and we're not seeing the necessary urgency to tackle the disease." Dr. Tido von Schoen-Angerer, Director, DWB’s Campaign for the Access to Essential Medicines