Assay for determining endogenous levels of analyte in vivo

1. A method for determining endogenous levels of stromal cell-derived factor 1 (SDF-1) isoforms in vivo comprising the steps of:

a. preparing a mixture of internal standard of SDF-1 isoforms, wherein the internal standard SDF-1 isoforms includes intact SDF-1 isoforms and NH 2 -terminal truncated SDF-1 isoforms, a plasma sample containing endogenous SDF-1, wherein the plasma sample is from a subject and wherein the endogenous levels of SDF-1 includes intact SDF-1 isoforms and NH 2 -terminal truncated SDF-1 isoforms and an immobilized antibody;

b. incubating the mixture;

c. separating out the immobilized antibody bound with SDF-1 isoforms;

d. releasing the SDF-1 isoforms from the immobilized antibody;

e. introducing the SDF-1 isoforms to a mass spectrometer; and

f. detecting the amount of endogenous in vivo levels of the SDF-1 isoforms wherein detection of the endogenous levels of SDF-1 includes detection of intact SDF-1 isoforms and NH 2 -terminal truncated SDF-1 isoforms.

2. The method of claim 1 wherein an intact SDF-1 isoform is SDF-1α.

3. The method of claim 1 wherein an intact SDF-1 isoform is SDF-β.

4. The method of claim 1 wherein the immobilized antibody is a SDF-1α antibody immobilized on a magnetic bead.

5. The method of claim 1 wherein the plasma sample is from a human.

6. The method of claim 1 wherein the plasma sample is from a mouse.

7. The method of claim 1 , wherein the linear range spans three orders of magnitude.

8. The method of claim 1 , wherein the amount of endogenous levels of the SDF-1 isoforms spans picamolar per liter to nanomolar per liter.

9. The method of claim 1 , wherein the method is capable of detecting SDF-1 at concentrations of less than 5 nmol/L in the sample.

10. The method of claim 1 , wherein the method is capable of detecting SDF-1 at concentrations of less than 2 nmol/L in the sample.

11. The method of claim 1 , wherein the method is capable of detecting SDF-1 at concentrations of less than 1 nmol/L in the sample.

12. A method for determining suppression of inactive stromal cell-derived factor (SDF-1) isoforms by dipeptidyl peptidase-4 (DPP-IV) inhibition in vivo comprising the steps of:

a. drawing a first plasma sample from a subject;

b. administering to the subject a DPP-IV inhibitor;

c. drawing a second sample from the subject;

d. assaying the first and second sample with the assay as described in claim 1 ; and

e. comparing the amount of intact SDF-1 isoforms from the DPP-IV cleaved NH 2 -terminal truncated SDF-1 isoforms.

13. The method of claim 12 , wherein the subject is human.

14. The method of claim 12 , wherein the subject is a mouse.

15. The method of claim 12 wherein an intact SDF-1 isoform is SDF-1α.

16. The method of claim 12 wherein an intact SDF-1 isoform is SDF-β.