Microfluidic protein crystallography techniques

• US 8,052,792 B2
• Filed: 05/15/2007
• Issued: 11/08/2011
• Est. Priority Date: 04/06/2001
• Status: Expired due to Fees

• Current Assignee: California Institute of Technology, Regents of the University of California (University of California)
• Original Assignee: California Institute of Technology, Regents of the University of California (University of California)
• Inventors: Quake, Stephen R., Hansen, Carl L., Berger, James M.
• Primary Examiner(s): KUNEMUND, ROBERT M
• Application Number:
  US11/748,838
• Publication Number:
  US 20070209574A1
• Time in Patent Office: 1,638 Days
• Field of Search: 117/68, 117/69, 117/70
• US Class Current: 117/68
• CPC Class Codes: B01D 9/00 Crystallisation crystallisation directly from the vapour phase B01D7/02; making single crystals C30B ; crystallisation as part of the Bayer process also classified in C01F7/14
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00355 peristaltic
• CPC Class Codes: B01J 2219/00378 Piezoelectric or ink jet dispensers
• CPC Class Codes: B01J 2219/00396 Membrane valves
• CPC Class Codes: B01J 2219/00398 in multiple arrangements
• CPC Class Codes: B01J 2219/00439 using micromirror arrays
• CPC Class Codes: B01J 2219/00599 Solution-phase processes
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00707 separated from the reactor apparatus
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01J 2219/00725 Peptides
• CPC Class Codes: B01J 2219/00756 Compositions, e.g. coatings, crystals, formulations
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2300/0867 Multiple inlets and one sample wells, e.g. mixing, dilution
• CPC Class Codes: B01L 2300/087 Multiple sequential chambers
• CPC Class Codes: B01L 2300/123 Flexible; Elastomeric
• CPC Class Codes: B01L 2400/0472 Diffusion
• CPC Class Codes: B01L 2400/0481 : squeezing of channels or chambers
• CPC Class Codes: B01L 2400/0655 : pinch valves
• CPC Class Codes: B01L 2400/084 : Passive control of flow resistance
• CPC Class Codes: B01L 3/06 : Crystallising dishes
• CPC Class Codes: B01L 3/5025 : for parallel transport of multiple samples
• CPC Class Codes: B01L 3/502738 : characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: B01L 3/502753 : characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation separation in general B01D; microapparatus for analysis using electrophoresis G01N27/44791; sample preparation G01N1/28
• CPC Class Codes: C30B 29/54 : Organic compounds
• CPC Class Codes: C30B 29/58 : Macromolecular compounds
• CPC Class Codes: C30B 35/00 : Apparatus not otherwise provided for, specially adapted for the growth, production or after-treatment of single crystals or of a homogeneous polycrystalline material with defined structure
• CPC Class Codes: C30B 7/00 : Single-crystal growth from solutions using solvents which are liquid at normal temperature, e.g. aqueous solutions from molten solvents C30B9/00; by normal or gradient freezing C30B11/00; under a protective fluid C30B27/00
• CPC Class Codes: C30B 7/14 : the crystallising materials being formed by chemical reactions in the solution
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: G01N 1/08 : involving an extracting tool, e.g. core bit
• CPC Class Codes: Y10T 117/1024 : for crystallization from liquid or supercritical state

Abstract

The present invention relates to microfluidic devices and methods facilitating the growth and analysis of crystallized materials such as proteins. In accordance with one embodiment, a crystal growth architecture is separated by a permeable membrane from an adjacent well having a much larger volume. The well may be configured to contain a fluid having an identity and concentration similar to the solvent and crystallizing agent employed in crystal growth, with diffusion across the membrane stabilizing that process. Alternatively, the well may be configured to contain a fluid having an identity calculated to affect the crystallization process. In accordance with the still other embodiment, the well may be configured to contain a material such as a cryo-protectant, which is useful in protecting the crystalline material once formed.

Claims

1. A crystal growth method comprising:

disposing a solution including a solvent and containing a crystal source material in a first chamber defined by an elastomer structure;

disposing a crystallizing agent in a second chamber defined by the elastomer structure;

disposing a material in a well defined by the elastomer structure adjacent to and separated from the first and second chambers by an elastomer membrane; and

placing the first and second chambers in fluid communication to alter a solubility of the crystal source material, such that a presence of the material affects formation of a crystal from the crystal source material.

2. The method of claim 1 wherein the material exhibits a defined permeability to water, such that the presence of the material alters a rate of diffusion of water from at least one of the first and second chambers.

3. The method of claim 2 wherein the material comprises an oil.

4. The method of claim 2 wherein a thickness of the material within the well regulates the rate of water diffusion.

5. The method of claim 1 wherein the elastomer is permeable to a component of the material, such that the material diffuses into at least one of the first and second chambers.

6. The method of claim 5 wherein the component comprises water.

7. The method of claim 5 wherein the material comprises a cryoprotectant.

8. The method of claim 5 wherein the material comprises the crystallizing agent.

9. The method of claim 5 wherein the material comprises the solvent.

10. The method of claim 5 wherein the component comprises an additive other than the solvent and the crystallizing agent.

11. The method of claim 10 wherein the component comprises a cross-linking reagent.

12. The method of claim 10 wherein the component comprises a ligand.

13. The method of claim 10 wherein the component comprises a small molecule drug.

14. The method of claim 5 wherein diffusion of the material occurs during crystal formation.

15. The method of claim 5 wherein diffusion of the material occurs subsequent to crystal formation.

16. The method of claim 5 wherein diffusion of the material occurs across a PDMS membrane having a thickness of about 500 □

m or less.

17. The method of claim 1 further comprising irradiating the elastomer to produce a diffraction pattern corresponding to the crystal.

18. The method of claim 17 further comprising physically separating a crystal-containing portion of the elastomer from the surrounding elastomer prior to the irradiation.

19. The method of claim 1 further comprising introducing a cryo-protectant to the crystal.

20. The method of claim 19 further comprising introducing a cross-linking reagent to the crystal.

21. The method of claim 1 further comprising introducing a cross-linking reagent to the crystal.

22. An apparatus for forming crystals, the apparatus comprising:

an elastomer structure having a Young'"'"'s modulus of between about 20 Pa and 1 GPa defining a first chamber in selective fluid communication with a second chamber, the first chamber configured to contain a solution containing a crystal material dissolved in a solvent, and the second chamber configured to contain a crystallizing agent, the first and second chambers separated from an adjacent well by a thin elastomer membrane.

23. The apparatus of claim 22 further comprising a substrate in contact with the elastomer structure to define the first and second chambers.

24. The apparatus of claim 22 wherein the substrate is planar and a volume of the first and second chambers is defined by a depth of a recess in the elastomer.

25. The apparatus of claim 22 wherein the substrate comprises a recess partially defining a volume of at least one of the first and second chambers.

26. The apparatus of claim 22 further comprising a cap structure overlying the well.

27. The apparatus of claim 22 wherein the first and second chambers are in selective fluid communication with one another through a flow channel controlled by a microfluidic valve.

28. The apparatus of claim 22 further comprising a material disposed in the well, the material selected from the group consisting of an oil and a solvent-containing species.

29. The apparatus of claim 28 wherein the solvent-containing species is selected from the group consisting of the solvent, the crystallizing agent, a cryo-protectant, a cross-linking reagent, a ligand, and an additive.

30. The apparatus of claim 22 wherein the first and second chambers define a total reaction volume of at least about 500 nL.

31. The apparatus of claim 22 wherein the elastomer structure having a Young'"'"'s modulus of between about 50 Pa and 10 MPa.

32. The apparatus, of claim 31 wherein the elastomer structure, having a Young'"'"'s modulus of between about 100 Pa and 1 MPa.