Method for screening crystallization conditions in solution crystal growth

• US 20030022384A1
• Filed: 05/30/2002
• Published: 01/30/2003
• Est. Priority Date: 04/06/1999
• Status: Active Grant

• Current Assignee: UAB Research Foundation (University of Alabama System)
• Original Assignee: UAB Research Foundation (University of Alabama System)
• Inventors: DeLucas, Lawrence James
• Granted Patent: US 7,247,490 B2
• Time in Patent Office: Days
• US Class Current: 436/86
• CPC Class Codes: B01D 9/0063 Control or regulation control per se G05
• CPC Class Codes: B01D 9/0072 Crystallisation in microfluidic devices
• CPC Class Codes: B01D 9/0077 Screening for crystallisation conditions or for crystal forms
• 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 2219/00313 the reactor vessels being formed by arrays of wells in blocks
• CPC Class Codes: B01J 2219/00369 in multiple or parallel arrangements
• CPC Class Codes: B01J 2219/00479 Means for mixing reactants or products in the reaction vessels
• CPC Class Codes: B01J 2219/00495 Means for heating or cooling the reaction vessels
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/00599 Solution-phase processes
• CPC Class Codes: B01J 2219/00693 Means for quality control
• CPC Class Codes: B01J 2219/00707 separated from the reactor apparatus
• CPC Class Codes: B01J 2219/00725 Peptides
• CPC Class Codes: B01J 2219/00756 Compositions, e.g. coatings, crystals, formulations
• CPC Class Codes: B01L 2300/041 Connecting closures to device or container
• CPC Class Codes: B01L 2300/0819 Microarrays; Biochips
• CPC Class Codes: B01L 3/06 Crystallising dishes
• CPC Class Codes: B01L 3/5085 for multiple samples, e.g. microtitration plates
• CPC Class Codes: C30B 29/58 Macromolecular compounds
• 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: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 50/08 : Liquid phase synthesis, i.e. wherein all library building blocks are in liquid phase or in solution during library creation; Particular methods of cleavage from the liquid support
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: G01N 33/00 : Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
• CPC Class Codes: G01N 33/68 : involving proteins, peptides or amino acids involving lipoproteins G01N33/92

Abstract

A method of screening protein crystal growth conditions with picogram to microgram amounts of protein in picoliter or nanoliter volumes is provided. A preferred method comprises a microarray with a plurality of micro-chambers in the microarray. A protein solution is placed into the micro-chambers by an automated dispensing mechanism. The protein crystal growth conditions of each of the micro-chambers is adjusted so that the protein crystal growth conditions in at least two of the micro-chambers differs. Crystallization of the protein solution in the micro-chambers is effected. Protein crystal growth in the micro-chambers is then observed.

Claims

1. A method of screening protein crystal growth conditions employing picogram, nanogram or to microgram amounts of protein comprising the steps of:

providing a microarray with a plurality of wells in said microarray;

accurately dispensing a volume of from about 0.001 nl to about 250 nl of a protein solution into said wells;

controlling the protein crystal growth conditions of each of said wells so that the protein crystal growth conditions in at least two of said micro-chambers differs; and

observing protein crystal growth or protein precipitation in said wells.

2. The method of claim 1 wherein said protein solution comprises a component selected from the group consisting of buffers, surface active agents, salts, alcohols, polyethylene glycol and mixtures thereof.

3. The method of claim 1 wherein said protein solution is buffered.

4. The method of claim 1 wherein controlling protein crystal growth comprises employing a precipitate solution that is in fluid communication with the microarray.

5. The method of claim 1 wherein controlling protein crystal growth comprises adding a precipitate solution to the microarray.

6. The method of claim 5 wherein said precipitate solution and said protein solution are in different wells in said microarray and said microarray has channels for fluid communication between a well comprising protein solution and a well comprising a precipitate solution.

7. The method of claim 6 wherein controlling protein crystal growth further comprises employing varying dimensions of the channels.

8. The method of claim 6 wherein said precipitate solution and said protein solution are in fluid communication via micro-channels.

9. The method of claim 5 wherein said precipitate solution and said protein solution are in fluid communication.

10. The method of claim 9 wherein fluid communication is by liquid-liquid diffusion of said precipitate solution and said protein solution.

11. The method of claim 9 wherein said precipitate solution has a lower vapor pressure than the protein solution and fluid communication is by vapor diffusion.

12. The method of claim 5 wherein the protein solution and precipitate solution are in the same well and said crystallization is effected by batch crystallization.

13. The method of claim 12 wherein said wells further comprise a buffer solution.

14. The method of claim 5 wherein the precipitate solution has a volume from about 0.001 nl to about 250 nl.

15. The method of claim 1 wherein protein crystal growth or protein precipitation is observed by microscopy.

16. The method of claim 15 wherein the microscopy is differential interference contrast microscopy.

17. The method of claim 1 wherein the protein solution is dispensed into said wells by fast solenoid dispensing.

18. A method of screening protein crystal growth conditions employing picogram to microgram amounts of protein comprising the steps of:

accurately dispensing a volume from about 0.001 nl to about 250 nl of a protein solution onto a platform;

controlling the protein crystal growth condition of the sample; and

observing a protein precipitate or protein crystals in the sample.

19. The method of claim 18 wherein the sample is accurately dispensed by fast solenoid dispensing.

20. The method of claim 18 wherein controlling the protein crystal growth condition comprises employing a protein precipitate solution.

21. The method of claim 20 wherein said precipitate solution has a lower vapor pressure than the protein solution and crystallization is effected by vapor diffusion.

22. The method of claim 20 wherein the protein solution and precipitate solution mixed together and said crystallization is effected by batch crystallization.

23. The method of claim 20 wherein crystallization is effected by liquid-liquid diffusion of said precipitate solution and said protein solution.

24. The method of claim 18 wherein the platform is a microarray.

25. The method of claim 18 wherein protein crystal growth or protein precipitation is observed by microscopy.

26. The method of claim 25 wherein the microscopy is differential interference contrast microscopy.

27. A microarray for screening protein crystal growth at nanogram or picogram protein amounts comprising:

a plurality of wells wherein said wells are adapted for holding volumes of protein solution from about 0.001 nl to about 250 nl;

and further wherein said well comprises a material that is minimally water absorbing; and

an optically clear path from said wells.

28. The microarray of claim 27 wherein said wells further comprise a material that is substantially hydrophobic.

29. The microarray of claim 27 further comprising a plurality of wells for holding a precipitate solution and wherein said microarray has channels for fluid communication between wells holding protein solution and wells holding precipitate solution.

30. The microarray of claim 29 wherein at least two channels have different dimensions.

31. The microarray of claim 27 wherein said wells of said microarray can be sealed to prevent evaporation from said wells.

32. A microarray for screening protein crystal growth at nanogram or picogram protein amounts comprising a plurality of wells wherein said wells are adapted for holding volumes of protein solution from about 0.001 nl 5 to about 250 nl.

33. The microarray of claim 32 wherein said wells comprise a material that is minimally water absorbing.

34. The microarray of claim 32 further comprising an optically clear path from said wells.

35. The microarray of claim 32 wherein said wells comprise a material that is substantially hydrophobic.

36. The microarray of claim 32 further comprising a plurality of wells for holding a precipitate solution and wherein said microarray has channels for fluid communication between wells holding protein solution and wells holding precipitate solution.

37. The microarray of claim 36 wherein at least two channels have different dimensions.

38. The microarray of claim 32 wherein said wells in said microarray can be sealed to prevent evaporation from said wells.