Bioinspired of Micro-fluidic systems Reach Symposium-2008

1. Bioinspired of Micro-fluidic systems Reach Symposium-2008 Shantanu Bhattacharya Assistant Professor Department of Mechanical Engineering Indian Institute of Technology Kanpur bhattacs@iitk.ac.in Tel: 0512-259-6056

2. The field of Bio-mimetics • 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.

3. 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

4. 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

5. Materials in Micro-fluidic devices • Silicon and microelectronic materials • Glass, Quartz Alternate Biochip materials • Polymers – Poly (dimethylsiloxane) (PDMS) – Poly (methyl methacrylate) (PMMA) – Teflon, etc. • Biological Entities – Cells, Proteins, DNA – Frontier of BioMEMS !

6. 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.

7. Device fabrication using polymers

8. Micro-channel Arrays using Controlled Etching Real image of micro-channels after swelling in solvent Ref: Sharma et. al., Science, 2007 1- Dimensional 2- Dimensional 3- Dimensional Cross-sectional View

9. 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. Bottom up Ref :Stephen D. Centuria, Microsystem Design, Kluwer Academic Publishers, Boston / Dordrecht / London Lectures, from NanoHUB, Purdue University, West Lafayette, Indiana

10. Content of presentation • Mimicking Biological architectures for certain engineering end goals. • Mimicking Biological principles for certain engineering end goals. • Micro-fluidics and Bio-sensing

11. Micro-fluidic Sample Preparator • 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

12. Microfluidic tectonics: A comprehensive construction platform for microfluidic systems • 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 Beebe et. al., 2000, PNAS, Vol. 97, pp. 13488-13493.

13. 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. 3 Kim et.al., NSTI-Nanotech 2005, Vol.1

14. Nanoscale DNA coulter Counter

15. Pore Shrinking and Shape Changing (after Thermal Oxidation, the oxide thickness is 50nm)

16. Nanopore channel sensors for characterization of dsDNA

17. Biochips driven by Bioinspired Microfluidics Bhattacharya et. al., JMEMS, 2007. Chang et. al., Biomedical Microdevices, 2003. Bhattacharya et. al., Lab chip, 2007, under review.

18. Lab on a chip for Viral detection

19. 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

20. Polymerase Chain Reaction

21. DNA translation in Agarose (Electrophoresis) I 1 +ve -ve II Sequential fluorescent images of DNA migration behavior in mediums: (a) Nanospehere (b) Agarose and (c) Control Buffer solution without nanosphere 1 +ve -ve [1] Nanospheres for DNA separation chips Mari 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 III

22. Equipment in a PCR laboratory DNA Extraction from tissue samples Imaging of Fluorescence Glove box for preparing the PCR Mix Gel electrophoresis of DNA PCR thermal Cycler

23. Lab on Chip Design for the Analyzer Non fluorescing reference channel for background subtraction Syringe for injecting PCR mix and sample Electrodes for Gel electrophoresis Micro channel filled with agarose gel Compressed air bottle LED Plan View Front Elevation View Spectrometer Reference Solid Core Waveguide for background subtraction Solid Core Waveguides placed along target DNA regions Lab-view Operated Solenoid valve Heaters for PCR DAQ system hooked to spectrometer will provide the spatial data for the differential intensities Optical Fibers from Assay Computer with DAQ card Spectrometer

24. 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. Pneumatic Chambers Inflow Fluid Channels Outflow Pumping Cycle Pumps in action Picture of the pumps

25. Peristaltic Pumps in action

26. Working PCR Chip for IBR isolates Amplified Extract from chip Amplification performed on .07 pg/ μl sample conc. Amplified Sample from Conventional M/c 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.

27. Capillary Electrophoresis: Sample and Capillary Loading 2 Basic Capillary Designs A B Sample loading sequence in Gel filled channels

28. Capillary Electrophoretic Chip Reduces Detection Time by a Factor of 40 1.5% agarose solution in microchannels 300 V for 50 secs 300 V for 25 secs 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 Conventional Electrophoresis Time= 35mins Requirement : Low voltage capillary electrophoresis system 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).

29. 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) Average particle size= 13.16nm, + 3.93nm

30. SEM/ TEM images of the doped gels TEM image of Platinum doped agarose Back scattered image (FESEM) Array image of platinum particles embeded in agarose Sizes: 2.5 microns 500 nm 500 nm Ack.: Lou Ross, Randy Tindel and Cheryl Jensen., EMC core EDS spectra of the Platinized gels

31. Enhanced DNA mobility Mobility Enhancement 2 times at 16V/cm Calculations done using the one dimensional mobility model µ = v/ E where , µ = mobility of the stain, v= Velocity (cm/ sec.), E= Electric Field (V/cm)

32. Cdi Rser Rs Zw Zw Dielectric Constant Enhancement due to nano-platinum Electrode spacing = 23microns Electrode width= 17microns Mobility = ε ε0 ζ / η [1] εhas approx. 2 times enhancement [1] Rieger T.H., “Electrochemistry”; Prentice Hall, inc., New Jersey, 1987 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).

33. 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.

34. ACKNOWLEDGEMENTS Collaborators and Advisors: • 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). 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)