Volatile matrices for matrix-assisted laser desorption/ionization mass spectrometry

• US 6,104,028 A
• Filed: 05/29/1998
• Issued: 08/15/2000
• Est. Priority Date: 05/29/1998
• Status: Expired due to Term

• Current Assignee: Agena Bioscience Inc. (Mesa Laboratories Inc.)
• Original Assignee: GeneTrace Systems Inc.
• Inventors: Lin, Hua, Hunter, Joanna M., Becker, Christopher H.
• Primary Examiner(s): NGUYEN, KIET TUAN
• Application Number:
  US09/086,993
• Time in Patent Office: 809 Days
• Field of Search: 250/288, 250/281, 250/282, 436/173
• US Class Current: 250/288
• CPC Class Codes: G01N 1/22 in the gaseous state specially adapted for biological material G01N33/497; measuring breath flow A61B5/087
• CPC Class Codes: H01J 49/0468 with means for heating or cooling the sample
• CPC Class Codes: H01J 49/164 Laser desorption/ionisation, e.g. matrix-assisted laser desorption/ionisation [MALDI] sample holders H01J49/0418

Abstract

A sample preparation method is disclosed for volatilization and mass spectrometric analysis of nonvolatile high molecular weight molecules. Photoabsorbing molecules having significant sublimation rates at room temperature under vacuum, and preferably containing hydroxy functionalities, are disclosed for use as matrices in matrix-assisted laser desorption/ionization mass spectrometry. The samples are typically cooled in the mass spectrometer to temperatures significantly below room temperature.

Claims

1. A method for determining the mass of a large organic molecule, said method comprising:

(a) contacting said large organic molecule with a photoabsorbing low-sublimation temperature matrix to produce a matrix;

molecule mixture;

(b) desorbing and ionizing said molecule; and

(c) determining the mass of said ionized large organic molecule by mass spectrometry.

2. The method of claim 1, wherein said mass spectrometry is performed using a cooled sample stage.

3. The method of claim 2, wherein said sample stage is cooled by liquid nitrogen.

4. The method of claim 3, wherein said sample stage is cooled to from about 170°

K. to about 190°

K.

5. The method of claim 4, wherein said sample stage is cooled to about 180°

K.

6. The method of claim 1, wherein said contacting is performed by dissolving said large organic molecule in a solution containing said matrix.

7. The method of claim 6, wherein said solution comprises a solvent.

8. The method of claim 7, wherein said solvent is organic.

9. The method of claim 7, wherein the solvent is water, methanol, ethanol, toluene, acetone, or acetonitrile.

10. The method of claim 7, wherein said solvent is evaporated prior to mass spectrometry.

11. The method of claim 10, wherein said solvent is evaporated at room temperature.

12. The method of claim 1, wherein said matrix:

molecule mixture is cooled to a vapor pressure between about 10^-5 Torr and about 10^-10 Torr prior to mass spectrometry.

13. The method of claim 1, wherein said matrix is further defined as a volatile, light-absorbing, hydroxy-bearing matrix.

14. The method of claim 1, wherein said matrix is selected from the group consisting of phenols, hydroxyquinolines, and hydroxynaphthalenes.

15. The method of claim 14, wherein said matrix is a phenol.

16. The method of claim 15, wherein said phenol is 4-nitrophenol.

17. The method of claim 14, wherein said matrix is a hydroxyquinoline.

18. The method of claim 17, wherein said hydroxyquinoline is 8-hydroxyquinoline.

19. The method of claim 1, wherein said large organic molecule is a polymer.

20. The method of claim 19, wherein said polymer is a biopolymer.

21. The method of claim 20, wherein said biopolymer is selected from the group consisting essentially of nucleic acid, polypeptide, peptide nucleic acid and oligosaccharide.

22. The method of claim 21, wherein said biopolymer is a nucleic acid.

23. The method of claim 22, wherein said nucleic acid is DNA.

24. The method of claim 22, wherein said nucleic acid is RNA.

25. The method of claim 1, wherein said large organic molecule has a mass of greater than about 1,000 Da.

26. The method of claim 25, wherein said large organic molecule has a mass of greater than about 27,000 Da.

27. The method of claim 26, wherein said large organic molecule has a mass of greater than about 250,000 Da.

28. The method of claim 27, wherein said large organic molecule has a mass of greater than about 315,000 Da.

29. The method of claim 1, wherein said matrix has a sublimation rate at room temperature of at least 0.1 μ

m·

min^-1 at a pressure of less than about 10^-5 Torr.

30. The method of claim 1, wherein said matrix has a sublimation rate at room temperature of from about 0.01 μ

m·

min^-1 to about 0.1 mm·

min^-1 at a pressure of less than about 10^-5 Torr.

31. The method of claim 1, wherein said matrix has an absorption coefficient greater than about 101·

cm^-1 ·

mol^-1.

32. The method of claim 1, wherein said desorbing comprises exposing said matrix:

molecule mixture to a source of energy to desorb said large organic molecule from the matrix.

33. The method of claim 32, wherein said source of energy is pulsed laser radiation.

34. The method of claim 33, wherein said pulsed laser radiation comprises applying an energy of about 20 kV followed by a pulse of energy of about 2.7 kV pulse.

35. The method of claim 34, wherein said pulse of energy comprises light having a wavelength of about 355 nm.

36. The method of claim 1, wherein said mass spectrometry comprises time-of-flight mass spectrometry.

37. The method of claim 36, wherein the mass of said ionized large organic molecule is determined by summing mass spectra over about 200 laser pulses.

38. A method for preparing a sample of large organic molecules for mass spectral analysis, said method comprising:

a) providing a solution comprising said large organic molecule to be analyzed, a volatile, light-absorbing hydroxy-bearing matrix composition, and a solvent; and

b) evaporating said solvent to provide a solid matrix material containing the molecule to be analyzed.

39. The method of claim 38, wherein said matrix composition is selected from the group consisting essentially of phenols, hydroxyquinolines, and hydroxynaphthalenes.

40. The method of claim 39, wherein said matrix composition is a phenol.

41. The method of claim 40, wherein said phenol is 4-nitrophenol.

42. The method of claim 39, wherein said matrix composition is a hydroxyquinoline.

43. The method of claim 42, wherein said hydroxyquinoline is 8-hydroxyquinoline.

44. The method of claim 38, wherein said large organic molecule is a polymer.

45. The method of claim 44, wherein said polymer is a biopolymer.

46. The method of claim 45, wherein said biopolymer is selected from the group consisting of nucleic acid, polypeptide, peptide nucleic acid and oligosaccharide.

47. The method of claim 46, wherein said biopolymer is a nucleic acid.

48. The method of claim 47, wherein said nucleic acid is DNA.

49. The method of claim 47, wherein said nucleic acid is RNA.

50. The method of claim 38, wherein said large organic molecule has a mass of greater than about 1,000 Da.

51. The method of claim 50, wherein said large organic molecule has a mass of greater than about 27,000 Da.

52. The method of claim 51, wherein said large organic molecule has a mass of greater than about 250,000 Da.

53. The method of claim 52, wherein said large organic molecule has a mass of greater than about 315,000 Da.

54. The method of claim 38, wherein said matrix has a sublimation rate at room temperature of at least 0.1 μ

m·

min^-1 at a pressure of less than about 10^-5 Torr.

55. The method of claim 38, wherein said matrix has an absorption coefficient greater than about 101·

cm^-1 ·

mol^-1.