Microfluidic Dialysis Protein Crystallization Jiang Huang GN Biosystems, Inc. March 26, 2009
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Transcript of Microfluidic Dialysis Protein Crystallization Jiang Huang GN Biosystems, Inc. March 26, 2009
Microfluidic Dialysis Protein Crystallization
Jiang HuangGN Biosystems, Inc.
March 26, 2009
Dialysis Protein Crystallization Method
Pro: scans a wide concentration range, the reagent composition can be easily altered during the course of the experiment.
Con: difficult to setup, protein consumption too high (5 to 350l/rxn), not HT compatible.
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Lysozyme
Glucoseisomerase
Catalase
Microfluidic Dialysis Plate - Design
open bottom microtiter plate
protein inlet film
dialysis membrane discs
microfluidic plate
adhesive sealing tape
protein inlet
vacuum port
dialysis membrane reagent well
Microfluidic Dialysis Plate - Design
adhesive film
Microfluidic Dialysis Plate Design
Top View
Dialysis chambers dimensions:
Screening plate:18nl per chamber (240m dia., 400m deep).
Optimization plate: 80nl per chamber (500m dia., 400m deep).
Growth plate: 1l per chamber (1.6mm dia., 400m deep)
Bottom View
Vacuum port
Dialysis chamber
Microfluidic channel
Protein port
Microfluidic Dialysis Plate – Sample Loading
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
Equipment and Accessories Needed
1. Vacuum pump or house vacuum with an ultimate vacuum ≤ 0.1mmHg*Air bubbles in every dialysis chamber will result due to insufficient vacuum
A List of Qualified Vacuum Pumps at under $2,000
Manufacture Model# Ultimate Vacuum
BOC/Edwards RV3 1×10-6mmHgBOC/Edwards EVA480-16-941 8×10-3mmHgWelch 1400B-01 1×10-4mmHgWelch 1399B-01 0.02mmHgBrinkmann V-500 0.01mmHgThermo-Electron 3178712 3.8×10-3mmHgThermo-Electron 3178707 1×10-4mmHg
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
via diameter: 1.2mm (570nl volume) via diameter: 0.3mm (20nl volume)crystal size: up to 500m long crystal size: up to 50m long
50mg/ml Lysozyme vs. HCS1 #10
Translation and Scale-up
screening optimization growthdia.=240m dia.=500m dia.=1.6mmdepth=400m depth=400m depth=400m
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Hampton Crystal Screen I
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Translation and Scale-up
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
FD Method Highlights
• Chemically compatible with commercial reagent kits
(materials used: PMMA, epoxy, dialysis membrane)
M. W. Toepke, D. J. Beebe, PDMS absorption of small molecules and consequences in microfluidic applications, Lab Chip, 2006, 6: 1484-1486
FD Method Highlights
• Protein consumption as low as 15nl per dialysis
chamber
• Fast set-up in as little as 4 minutes for 96 or 384
dialysis chambers
• Low capital equipment costs to begin running
experiments
• High-throughput compatibility with standard dispensing
robotics
• Easy translation and scale-up designs
• Chemically compatible with commercial reagent kits
• The dialysis membrane can be conveniently to allow
easy loop access for crystal manipulation
protein inlet
vacuum port
dialysis membrane reagent well
Microfluidic Dialysis Plate - Design
adhesive film