1	Multilayer Soft Lithography MSL for Life Sciences LRIG Mid Atlantic Bridgewater, NJ September 5 Rodney Turner
2	A Transistor for Fluidics
3	Successful Integration for Biology What was needed: Reliable, easy to fabricate valves and pumps reduce device footprints Materials matched to task bio-compatibility Scalable control scheme allow high level of integration
4	Multilayer Soft Lithography
5	MSL - principle
6	Multi-layer Soft Lithography – the MSL™ process
7	Fluidigm Micro-Pump
8	Fluidigm Micro-Mixer
9	What Fluidigm Has Done with MSL Taken full advantage of rapid prototyping capability “Gandalf” feasibility test: over 100 designs to chips in 4 months FluidArchitect™ Software Eliminate prototype development as bottleneck
10	What Fluidigm Has Done with MSL Enable four key life science applications Tailor-made miniaturization for protein crystallization: enabled by over 400 valves. High density PCR microprocessors for genotyping and gene expression: 1000s to 10,000s of nanoliter reaction volumes per chip. Unique PDMS gas permeability enables multiple live cell handling capabilities: culture, perfuse, lyse and analyze in a single chip Miniaturized, heterogeneous protein assays in self-programmable microprocessors: hundreds of valves and pumps per chip.
11	Fluidigm Protein Crystallizer: BENEFITS Parallel capacity: 144 experiments in one microprocessor. Sample reduction: 2 orders of magnitude reduction in sample input amount. Automation: Reduced sample handling and pipetting with improved reproducibility. New crystallization conditions: Offers potential to discover crystallization conditions for proteins that have not been crystallized before.
12	Protein Crystallization Starter Kit™ Ten 48-reagent microprocessors Ten microprocessor carriers Two microprocessor control boxes Two 8-channel manifolds Two 5-valve manifolds Detailed Protocol Control reagent set: Thaumatin crystallization
13	Crystallization Microprocessor: Blind Filling enabled by PDMS
14	Thaumatin Crystals in Microprocessor
15	Validation Experiments^*
16	Glucose Isomerase Results
17	Real World Test: Fluidigm/Caltech/UC Berkeley collaboration A Bacterial RNase No crystals after a set of full commercial screens Crystals discovered in microprocessors Reproduced by sitting drop based on results from microprocessors Bacterial Primase Conventional screens gave only “promising” precipitate Microprocessors gave clear crystals in initial screens Conventional conditions found faster Topoisomerase ATPase New conditions discovered in microprocessors Reproduced in conventional techniques Bacterial Ribosome Proteins Also crystallized (data proprietary)
18	Microprocessor Carrier for Easy Loading and Inspection
19	Fluidigm Protein Crystallizer: Future work Co-crystallization Crystal recovery chip Observe compound-induced physical changes to crystals
20	Why Microfluidics and Genetic Analysis? 10,000 quantitative PCR assays in a single microprocessor. Single loading step for 10,000 reactions. Decrease consumption of amplification reagents 5000-fold.
21	Chip Design: Sniper
22	Well-populated solution space
23	Single Sample Load Blind Fill Capability
24	Signal Strength Fluidigm PCR cocktail à good signal strength in microfluidic format
25	Digital template dilution Stepwise reduction in the amount of DNA gives the expected PCR signature:
26	Reagent Spotting and Delivery 1 nl reaction volumes
27	Nucleic Acid Microprocessor Benefits Harness the power of quantitative PCR 1 copy per 1,000 cells 5 orders-of-magnitude linear dynamic range Conduct 10,000 PCR reactions with a single loading step Reduce threshold cost for PCR analysis Reducing reaction volumes 5000x
28	Next Generation Biology Key Features Miniaturization High Content Analysis Integration
29	A Modular Approach
30	Reagent Perfusion Principle Focused Delivery Mixing is diffusion limited Precise control over reagent addition
31	Single Cell Capture & Perfusion Trypan blue staining pre and post methanol perfusion 100 millisecond perfusion time!!!
32	Multiple Cell Capture PBLs stained with Acridine Orange
33	The Live-Cell Science Microprocessor
34	Cell Catch and Culture
35	Cell Culture Time Lapse
36	Size Filtration
37	The Protein Microprocessor Enable immunoassay profiles in miniaturized format Automated approach lowers cost and improves data quality Integrate with miniaturized sample preparation capability
38	The Microprocessor Fluidic Network Reagent and sample are delivered sequentially to a matrix of intersecting channels bounded by isolation valves A unique assay occurs on the substrate at each channel intersection Fully automated delivery of samples, reagents, blocks and washes Processors conform to microscope slide dimensions Scan in standard array scanners
39	Flexible Surface Chemistry
40	Principle of Operation
41	Processor Fluidics
42	Quantitative Resolution and Sensitivity
43	Integration of Cell Handling and Protein Processing
44	Fluidigm: The Opportunity to Access existing programs Protein Crystallization chips ready to use Your genotyping content for Nucleic Acid Microprocessor Explore New Capabilities in Life Sciences Integration of Genome, Proteome and Live-cell studies on a single platform Rapid Design Prototyping
45	Fluidigm: Contact www.fluidigm.com Rodney Turner VP Business Development rodney.turner@fluidigm.com Kristin Spataro Manager, Business Development kristin.spataro@fluidigm.com