Bioinspired of Micro-fluidic systems Reach Symposium-2008 Shantanu Bhattacharya Assistant Professor...
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Transcript of Bioinspired of Micro-fluidic systems Reach Symposium-2008 Shantanu Bhattacharya Assistant Professor...
Bioinspired of Micro-fluidic systems Reach Symposium-2008
Shantanu BhattacharyaAssistant Professor
Department of Mechanical EngineeringIndian Institute of Technology Kanpur
[email protected]: 0512-259-6056
• Quite often Nature works at Maximum Achievement at minimum effort level.
• The field of Bio-mimetics is the abstraction of a good design from nature.
• The field of Bio-mimetics started from Giovanni Borelli’s seminal De Motu Animalum of 1680.
The field of Bio-mimetics
Blood Capillaries and Micro-fluidics•The capillaries are the smallest blood vessels that distribute oxygenated blood.
•Inspired by the micro-circulation inside the capillaries and its uses one can think of Micro-fluidics
Properties of Micro flows
•Surface effects become prominent with high surface area to volume ratio.
•Low thermal mass and high heat transfer.
•Low value of Reynolds number and thus laminar flows which only result in diffusional mixing.
•Re is usually less than 100 and often less than 0.1 in micro-devices
Micro-fluidics
Materials in Micro-fluidic devices• Silicon and microelectronic materials• Glass, QuartzAlternate Biochip materials• Polymers– Poly (dimethylsiloxane) (PDMS)– Poly (methyl methacrylate) (PMMA)– Teflon, etc.• Biological Entities– Cells, Proteins, DNA– Frontier of BioMEMS !
Microfluidic device fabrication in Silicon
NEMS/ MEMS silicon fabrication•Conventional and new semiconductor manufacturing techniques are used.
•Etching, Deposition, Photolithography, Oxidation, Epitaxy etc.
•Deep RIE, Thick plating etc.Bulk and surface micromachining.
Micro-channel Arrays using Controlled
Etching
1- Dimensional 2- Dimensional 3- Dimensional Cross-sectional View
Real image of micro-channels
after swelling in solvent
Ref: Sharma et. al., Science, 2007
Micro-fluidics and Microsystems
Relationship with the Biological world
•Systems made up of very small components.(micron to nanometer scale)
•Relatively high applicability to the field of life science, biotechnology and medicine.
•That’s why they scale with some of the biological entities.
•Focus of micro-system research is shifting to micro fluidic systems.
Ref :Stephen D. Centuria, Microsystem Design, Kluwer Academic Publishers, Boston / Dordrecht / London
Lectures, from NanoHUB, Purdue University, West Lafayette, Indiana
Bottom up
Content of presentation
• Mimicking Biological architectures for certain engineering end goals.
• Mimicking Biological principles for certain engineering end goals.
• Micro-fluidics and Bio-sensing
•A micro-separation device is realized
•Whole blood enters the device through a 70 microns channel.
•Margination happens and leukocyte distribution is affected .
Bitensky et.al., 2005, Anal. Chem. 77, 933-937
Micro-fluidic Sample Preparator
Microfluidic tectonics: A comprehensive construction platform for microfluidic systems
Beebe et. al., 2000, PNAS, Vol. 97, pp. 13488-13493.
•There are a lot of passive valves in our veins , allowing the fluid flow in only one direction.
•Hydrogel is used to realize a valve which swells and de-swells in different pH’s
Characteristics of bacterial pumps in microfluidic systems
•The growth of flagellum in flagellated bacteria like E. Coli or Serratia Marcescens is a function of glucose conc.
•Flagellated bacteria are used in microchannels to paddle fluids at various flow rates.
Fig. 1 Fig. 2 Fig. 3Kim et.al., NSTI-Nanotech 2005, Vol.1
Biochips driven by Bioinspired Microfluidics
Chang et. al., Biomedical Microdevices, 2003.
Bhattacharya et. al., JMEMS, 2007.
Bhattacharya et. al., Lab chip, 2007, under review.
Lab on chip for daignostics of Infectious Bovine Rhinotracheitis
• Annual losses due to the bovine viral disease IBR to the Beef industry stands at US$ 10-40 million per million animals (Bennett & Done, 1992,Harkness, 1997, Houe et al., 2003b). or $560million per annum. http://www.livestock.novartis.com/pdf/Arsenal_BVD_KnowlEdge.pdf
• Originally recognized as a respiratory disease in swine herds in 1991.
• Mechanism of transmission are mainly confinement particularly in feedlots. The disease is rapidly spread to new arrivals for already infected species.
• Field diagnosis is extremely important.Detection is carried out using PCR based assay in laboratories which is
time consuming.
Ref: Infectious Porcine Diseases, L.R. Sprott and S. Wiske, Agricultural communications, 2002
DNA translation in Agarose (Electrophoresis)
I
II
-ve+ve
III
-ve+ve
Sequential fluorescent images of DNA migration behavior in mediums: (a) Nanospehere (b) Agarose and (c) Control Buffer solution without nanosphere 1
1
[1] Nanospheres for DNA separation chipsMari Tabuchi1, 5, 6, Masanori Ueda1, 5, Noritada Kaji1, 5, Yuichi Yamasaki2, 5, Yukio Nagasaki3, 5, Kenichi Yoshikawa4, 5, Kazunori Kataoka2, 5 & Yoshinobu Baba1, 5, 6, 7 , NATURE
Equipment in a PCR laboratory
Glove box for preparing the PCR Mix
PCR thermal Cycler
Gel electrophoresis of DNA
Imaging of Fluorescence DNA Extraction from
tissue samples
DAQ system hooked to spectrometer will
provide the spatial data for the differential
intensities
Computer with DAQ card
Lab-view Operated Solenoid
valve
Spectrometer
Electrodes for Gel
electrophoresis
Heaters for PCR Optical Fibers
from Assay
Micro channel filled with agarose
gel
Reference Solid Core Waveguide for
background subtraction
Non fluorescing reference channel for background
subtraction
LED
Solid Core Waveguides placed along target DNA regions
Compressed air bottle
Plan View
Spectrometer
Syringe for injecting PCR mix
and sample
Front Elevation View
Lab on Chip Design for the Analyzer
Peristaltic Micro-pumps for fluid transport•Peristalsis is the motion of fluid in channels through a traveling contractile.
•This effect has been successfully utilized for the control of fluid motion.
•Pumping rates in the range of 10~12 microliters at pumping frequency of 10 Hz. has been attained.
•The pumps are 3 layered devices fabricated using Glass and PDMS and are energized by an offchip compressed nitrogen supply regulated thru labview.
Outflow
Inflow
Pneumatic Chambers
Fluid Channels
Pumping Cycle Pumps in action Picture of the pumps
Working PCR Chip for IBR isolates
0 2000 4000 6000 800020
30
40
50
60
70
80
90
100
110
Te
mp
era
ture
(d
eg
.)
Time (secs)
Labview output of temperature vs. time of IBR cycle
4000 4020 4040 4060 4080 4100 4120 4140 4160 4180 420020
30
40
50
60
70
80
90
100
110
Tem
per
atu
re (
deg
.)
Time (secs)
One full thermal cycle
Amplified Extract from chip
Amplified Sample from Conventional M/c
Amplification performed on .07 pg/ μl sample conc.
Ref: “Optimization of design and fabrication process for realization of a PDMS-Silicon DNA amplification chip”, by Shantanu Bhattacharya, Venumadhav Korampally, Yuanfang Gao, Maslina Othman, Sheila A. Grant, Steven B. Klieboeker, Keshab Gangopadhyay, Shubhra Gangopadhyay”, Journal of Microelectromechanical systems,Vol.99, pp.1-10, 2007.
Conventional system
130mins.
On chip System 15mins.
Capillary Electrophoresis: Sample and Capillary Loading
2 Basic Capillary Designs
Sample loading sequence in Gel filled channels
A B
Capillary Electrophoretic Chip Reduces Detection Time by a Factor of 40
300 V for 25 secs 300 V for 50 secs
1.5% agarose solution in microchannels
Requirement : Low voltage capillary electrophoresis system
Conventional Electrophoresis Time= 35mins
DNA ladder Trial: 100-1000 bp movement in an Agarose capillary.
Mobility (μ) = 9.101E-4 cm2/ Vsec .
Velocity of the stain=.078 cm/sec
Electric field = 85.7 V/cm
Ref: Bhattacharya, S., Gangopadhyay K., Gangopadhyay, S., Sharp, P.R., “A low voltage capillary electrophoresis system using platinum doped agarose gels”, (Manuscript to be submitted to Biosensors and bioelectronics).
Low Voltage Electrophoresis by Applying Nanotechnology
Platinum nano-particles made in situ
Potassium Chloro-Platinate is reduced by sodium boro-hydride after coating with a monolayer of Mercapto-Succinic acid in a Schlenk line in inert Argon environment. (2 conc. of solution used are 11.6mM and 23.2mM)
3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 220
20
40
60
80
100
120
140
Pa
rtic
le C
ou
nt
Particle diameter (nm)
Particle size distribution (nm) based on a total of 550 particles
Average particle size= 13.16nm, + 3.93nm
SEM/ TEM images of the doped gels
EDS spectra of the Platinized gels
Array image of platinum particles embeded in agarose
Sizes:
2.5 microns
500 nm
500 nm
Back scattered image (FESEM)TEM image of Platinum doped agarose
Ack.: Lou Ross, Randy Tindel and Cheryl Jensen., EMC core
Enhanced DNA mobility
4 6 8 10 12 14 160.00006
0.00008
0.00010
0.00012
0.00014
0.00016
0.00018
Mob
ility
(cm
2/v.
sec)
Electric field (V/cm)
Mobility vs. electric field for plain agarose Mobility vs. electric field for doped agarose
Calculations done using the one dimensional mobility modelµ = v/ E
where , µ = mobility of the stain, v= Velocity (cm/ sec.), E= Electric Field (V/cm)
Mobility Enhancement 2 times at 16V/cm
Dielectric Constant Enhancement due to nano-platinum
Mobility = ε ε0 ζ / η [1]
ε has approx. 2 times enhancement
Ref: Bhattacharya, S., Chanda, N., Grant S.A., Gangopadhyay K., Gangopadhyay, S., Sharp, P.R., “High conductivity agarose nano-platinum composites”, (Manuscript under review in Analytical Chemistry).
[1] Rieger T.H., “Electrochemistry”; Prentice Hall, inc., New Jersey, 1987
Electrode spacing
= 23microns
Electrode width= 17microns
Rs
Cdi
ZwZw
Rser
I II III0
500
1000
1500
2000
2500
Die
lect
ric
cap
aci
tan
ce (
pF
)
Type of Material
I- Plain Agarose II- Agarose doped with 5.8mM Pt. Hydrosol III- Agarose doped with 11.6mM Pt. Hydrosol
Summary and conclusions
• Bio-inspired Micro-fluidic technology is widely applied for biomimetics and biosensing.
• Lab-on-Chip is a direct spinoff of this technology and is used for providing point of care diagnostics.
• There is huge market potential for these technologies for the numerous applications.
ACKNOWLEDGEMENTS
• Dr. Sheila Grant, Dr. Shubhra Gangopadhyay , Dr. Keshab Gangopadhyay, Dr. Steve Klieboeker, Dr. Lela Riley, Dr. Xudong Fan (University of Missouri, Columbia).
• Dr. Rashid Bashir, Dr. Arun Bhunia, Dr. Michael Ladisch. (Purdue University, Indiana).
• Dr. P. Panigrahi, Dr. Bikram Basu, Dr. Bishakh Bhattacharya (IIT- Kanpur).
Collaborators and Advisors:
Funding Agencies:
•NSF (Curriculum Research Curriculum Development).•NPB (National Pork Board).•NIH (Mutant Mouse).•USDA (Center for food safety engineering).•Initiation Grant (IIT-Kanpur, DORD)