METHOD OF DETERMINING A PROTEIN IN A SAMPLE

1 . A method of determining a protein in a sample, comprising the steps of:

(a) providing an aptamer, which is functionalised at the 5′-end with a labeling reagent comprising a photo-activatable crosslinking moiety, a labeling moiety, and a linker moiety connecting the labeling reagent to the 5′-end of the aptamer;

(b) incubating a sample comprising an aptamer target protein with the functionalised aptamer of step (a);

(c) irradiating the sample with ultraviolet light for crosslinking the aptamer via the photoactivatable crosslinking moiety to the protein; and

(d) determining the protein via the labeling moiety.

2 . The method according to claim 1 , wherein the photo-activatable crosslinking moiety is selected from the group comprising aryl azides (phenyl azide), diazirine derivatives such as 3-trifluoromethyl-3-phenyl-diazirine (TPD), and benzophenone derivatives.

3 . The method according to claim 1 , wherein the labeling moiety is selected from the group comprising biotin and desthiobiotin.

4 . The method according to claim 1 , wherein the linker moiety provides a spacer arm length of about 6 Å to 45 Å, preferably about 20 Å to 40 Å, between the 5′-end of the aptamer and the crosslinking moiety.

5 . The method according to claim 1 , wherein the labeling reagent is Sulfosuccinimidyl-2-[6-(biotinamido)-2-(p-azidobenzamido)hexanoamido]ethyl-1,3′-dithiopropionate.

6 . The method according to claim 1 , wherein the method comprises incubating a biotin-labeled cross-linked aptamer-protein complex formed in step (c) with avidin-, neutravidin-, or streptavidin-coated magnetic beads.

7 . The method according to claim 1 , wherein step (d) includes determining a protein in a cell, or in an exosome, or in a circulating microvesicle.

8 . The method according to claim 1 , wherein step (d) includes determining a protein on the surface of a cell, or an exosome, or a circulating microvesicle.

9 . Use of the method according to claim 1 , for the identification of aptamer target proteins, particularly of aptamer target proteins in the proteome of cells, tissue or organisms, particularly for proteins of exosomes or circulating microvesicles.

10 . Use of the method according to claim 1 , for cell surface labelling.

11 . Use of the method according to claim 6 for magnetic cell sorting.