Nanobioprocessing

1. Nanobioprocessing Larry P. Walker, Ph.D. Professor, Department of Biological and Environmental Engineering Cornell University

2. Agricultural and Environmental Bioprocess Engineering Research • Molecular mechanisms of polysaccharide degrading enzymes, • Solid state microbial processes and molecular ecology, • Mechanisms and kinetics of metal uptake and sequestration in plants, • Molecular filtration and analysis of biomolecules

3. Nanobiotechnology Center (NBTC) Cornell University “The Nanobiotechnology Center (NBTC) was established in January 2000 as a Science & Technology Center, with core funding from the National Science Foundation. Nanobiotechnology is an emerging area of scientific and technological opportunity that integrates nano/microfabrication and biosystems to the benefit of both.” www.nbtc.cornell.edu

4. Nanobiotechnology Center (NBTC) Cornell University “Nanobiotechnology is beginning to generate substantial new insights into how biological systems function, and likewise, nanobiotechnology will lead to the design of entirely new classes of micro- and nanofabricated devices and systems.”

5. Nanobiotechnology Center (NBTC)Cornell University • Cornell University • Clark Atlanta University • Howard University • Princeton University • Oregon Health Sciences University • Wadsworth Center – New York Department of Public Health

6. Nanobiotechnology Center (NBTC)Cornell University • Cellular Devices (David Lawrence, Wadsworth Center)– studies of engineered devices for separating and analyzing individual cells. • Nanoscale Cell Biology(Manfred Lindau, Cornell University) - micro- and nanofabricated tools to understand and utilize cellular responses on the nanoscale. • Biomolecular Dynamics(Michael Koonce, Wadsworth Center) – exploration and utilization of molecular motion and mechanical properties

7. Nanobiotechnology Center (NBTC)Cornell University • Cell-Surface Interactions (Harvey Hoch, Cornell University) – Investigating the response and control of cellular responses on structured surfaces • Biomolecular Devices and Analysis (Larry Walker, Cornell University) - investigation new approaches to high speed and highly sensitivity analysis of biomolecules. • Nanoscale Biomaterials (Dotse Sogah, Cornell University) – study of materials engineered at the nanoscale for new properties that can be exploited in nanobiotechnolgy.

8. Biomolecular Devices and Analysis Major research and development activities in the life sciences has generated the need for materials, methods, and devices for sorting, separating, and analyzing proteins, DNA and other biomolecules.

9. Membranes and Packings Semipermeable Membranes and Equilibrium Dialysis Michael Spencer, Lori Lepak, Jim Turner, Michele Caggana Fabrication of Integratable Semipermeable Membranes Jim Turner, Peter Russo, David Martin, David Lawrence, Michele Caggana, Bill Shain, Michael Isaacson, and Michael Spencer Porous Block Copolymers For Microfluidic Separation Chris Ober &Mingqi Li High Resolution Protein Purification using Block Copolymer Derived Mesoporous Silica-type Materials Uli Wiesner, Phong Du, Caroline Corner, &Larry Walker

10. Micro and Nanofluidic devices Micro and Nanofluidics for Molecular Separation and Detection Harold Craighead, John Henion, David Czaplewski, Yanou Yang & Jun Kameoka Multidimensional Microscale Separations of Cerebrospinal Fluid Proteins Kelvin Lee, Michael Shuler, Jim Engstrom, Bob Austin, Chen Li & Jinpian Diao Nanostructures for Enzyme Transport and Accessibility Assessment Larry Walker, Harold Craighead, Jennifer Guisado & Tina Jeoh

11. Biomolecular Devices and Analysis Some Common Activities: • Lithography • Chemical synthesis • Spinning of molecules • Assessing morphology • Assessing monodispersity • Assessing transport mechanisms •  Measurement & Analysis

12. Biomolecular Devices and Analysis • Biocompatibility, • Working with small volumes: pico- to micro-liters, • Sieving and sorting at the scale of 5, 10, 15 and 50 nm, • Monodispersity – tight control over pore size distribution and shape

13. Spun Organic membrane Si3N4 Si Example: Membrane Technology Spin Coating of Cellulose And Collagen onto silicon surfaces to create porous membranes

14. 4 layers collagen:No methyl orange diffuses in 10 hours (up to 48 hours!) 1 layer collagen:Methyl orange (MW 327) diffuses through in 10 min Fabrication of Integratable Semipermeable Membranes – M. Spencer 400 nm thick Collagen Pores 0-100nm Dyes of 270 – 390 Da

15. 400V/cm (c) (b) 12 hrs Block Co-polymersUli Wiesner & Chris Ober •  excellent biocompatibility of silica • easily accessible pore sizes from 8-50 nm • narrow pore size distributions Chemical Synthesis Photo Lithography

16. Nanostructures for Enzyme Transport and Accessibility • Studying cellulase binding and catalysis as a function of molecular diameter. • Study synergism in cellulase/ cellulose reaction systems.

17. Channel design & fabrication Make inlets and outlets Glass-Glass bonding Attach peripheral components New 2 stage bonding approach developed to permit alignment and bonding. Flow chart of the device fabrication process 5 μm Double-T Structure GMA-co-TRIM polymer Channel with affinity separation bed Multidimensional Microscale Separations ProteinsK. Lee

18. Embossed Top Plastic Chip Embossed Fluidic Channel Reservoir hole Emitter film Bonding Bottom Plastic Chip Micro and Nanofluidics for Molecular Separation and Detection – H. Craighead Electrospray Ionization

19. (a) (a) Triangle Nozzle 50m Berberine:FW=336 (b) (b) Taylor cone Plume Liquid jet 50m Application to Mass Spectrometry Analysis

20. Nanobiotechnology • Advanced Bioreactors • Advanced Energy Systems • Controlled Environment Agriculture Systems • Genomics • Proteomics • Metabolic Engineering • Protein Engineering NBTC At the Interface between Biology and Engineering

21. We are having fun!