Methods and compositions for detecting and isolating phosphorylated molecules using hydrated metal oxides
View Patent ↗The invention provides methods for detecting and isolating phosphomolecules using phosphoaffinity materials that comprise a hydrated metal oxide. In an embodiment, a method for detecting a phosphomolecule in a sample involves (a) contacting a sample with a phosphoaffinity material comprising a hydrated metal oxide, under conditions wherein a phosphomolecule is capable of binding to the phosphoaffinity material to form a phosphomolecule-phosphoaffinity material complex, and (b) detecting formation of a phosphomolecule-phosphoaffinity material complex, thereby detecting a phosphomolecule in the sample. In another embodiment, a method for isolating a phosphomolecule from a sample involves (a) contacting a sample with a phosphoaffinity material comprising a hydrated metal oxide, under conditions wherein a phosphomolecule is capable of binding to the phosphoaffinity material to form a phosphomolecule-phosphoaffinity material complex, wherein the hydrated metal oxide comprises yttrium, and (b) separating the phosphomolecule-phosphoaffinity material complex from the sample, thereby isolating the phosphomolecule from the sample.
1. A method for detecting a phosphomacromolecule in a sample comprising:
(a) contacting the sample with a phosphoaffinity material comprising titanium dioxide, under conditions wherein the phosphomacromolecule is capable of binding to the phosphoaffinity material to form a phosphomacromolecule-phosphoaffinity material complex, wherein the phosphoaffinity material further comprises a support and wherein the support is selected from the group consisting of a particle, bead, gel, matrix, membrane, filter, fiber, sheet, mesh, frit, resin, sample vessel, column, pipette tip, slide channel, and MALDI-TOF plate; and
(b) detecting formation of the phosphomacromolecule-phosphoaffinity material complex, thereby detecting the phosphomacromolecule in the sample.
2. The method of claim 1 , wherein the detecting comprises measuring binding between the phosphomacromolecule and the phosphoaffinity material.
3. The method of claim 2 , wherein the measuring comprises measuring a physiochemical property of the phosphomacromolecule, the phosphoaffinity material, or both, wherein the physiochemical property is selected from the group consisting of absorbance, transmission, mass measurement, fluorescence intensity, fluorescence polarization, fluorescence resonance energy transfer, time-resolved fluorescence, resonance light scattering, surface-enhanced Raman scattering, electron paramagnetic resonance, refractive index absorbance, nuclear magnetic resonance, surface Plasmon resonance, refractive index changes, and combinations thereof.
4. The method of claim 1 , wherein the detecting comprises detecting the phosphoaffinity material portion of the phosphomacromolecule-phosphoaffinity material complex.
5. The method of claim 4 , wherein the titanium dioxide of the phosphoaffinity material portion is detected.
6. The method of claim 5 , wherein the titanium dioxide is detected using mass spectrometry.
7. The method of claim 6 , wherein the mass spectrometry is inductively-coupled plasma mass spectrometry.
8. The method of claim 1 , further comprising labeling the phosphoaffinity material with a detectable tag prior to detecting.
9. The method of claim 8 , wherein the detectable tag is a fluorescent moiety.
10. The method of claim 1 , wherein the titanium dioxide is labeled with a fluorescent moiety.
11. The method of claim 1 , wherein the detecting comprises detecting the phosphomacromolecule portion of the phosphomacromolecule-phosphoaffinity material complex.
12. The method of claim 11 , wherein the phosphomacromolecule portion is detected by inductively-coupled plasma mass spectrometry.
13. The method of claim 1 , wherein the detecting comprises detecting fluorescence resonance energy transfer.
14. The method of claim 13 , wherein fluorescence resonance energy transfer occurs between a fluorescent tag on the phosphomacromolecule and a fluorescent tag on the phosphoaffinity material.
15. The method of claim 1 , wherein the titanium dioxide is in particle form.
16. The method of claim 1 , wherein the support comprises an inorganic material.
17. The method of claim 1 , wherein the support is a particle.
18. The method of claim 17 , wherein the particle comprises a colloidal metal.
19. The method of claim 1 , wherein the support comprises an organic material.
20. The method of claim 1 , wherein the support is a sheet.
21. The method of claim 20 , wherein the sheet comprises cellulose.
22. The method of claim 1 , wherein the support comprises a detectable tag.
23. The method of claim 1 , wherein the phosphomacromolecule is a phosphorylated polypeptide.
24. The method of claim 1 , wherein the sample comprises a support.
25. The method of claim 1 , wherein the sample comprises a detectable tag.
26. The method of claim 1 , further comprising:
(c) isolating the phosphomacromolecule-phosphoaffinity material complex from the sample, thereby isolating the phosphomacromolecule from the sample.
27. The method of claim 26 , further comprising separating the phosphomacromolecule from the phosphomacromolecule-phosphoaffinity material complex.
28. The method of claim 1 , wherein the phosphomacromolecule is a phosphorylated polynucleotide.
29. The method of claim 1 , wherein the phosphomacromolecule is a phosphorylated lipid.
30. The method of claim 1 , wherein the phosphomacromolecule is a phosphorylated carbohydrate.
31. The method of claim 1 , wherein the support is a magnetic particle.
32. The method of claim 1 , wherein the support is a bead.
33. The method of claim 1 , wherein the support is a magnetic bead.