Method for generating aptamers with improved off-rates
The present disclosure describes improved SELEX methods for producing aptamers that are capable of binding to target molecules and improved photoSELEX methods for producing photoreactive aptamers that are capable of both binding and covalently crosslinking to target molecules. Specifically, the present disclosure describes methods for producing aptamers and photoaptamers having slower dissociation rate constants than are obtained using prior SELEX and photoSELEX methods. The disclosure further describes aptamers and photoaptamers having slower dissociation rate constants than those obtained using prior methods. In addition, the disclosure describes aptamer constructs that include a variety of functionalities, including a cleavable element, a detection element, and a capture or immobilization element.
1. A mixture comprising HSA, casein and prothrombin.
2. The mixture of claim 1 , wherein the mixture is a buffer solution.
3. The mixture of claim 2 , wherein the pH of the buffer solution is about 7.5.
4. The mixture of claim 1 , wherein HSA is at about 0.1%.
5. The mixture of claim 1 , wherein casein is at about 10 μM.
6. The mixture of claim 1 , wherein prothrombin is at about 10 μM.
7. The mixture of claim 1 further comprising a nucleic acid molecule.
8. The mixture of claim 7 , wherein the nucleic acid molecule is an aptamer.
9. A formulation comprising HEPES, NaCl, KCl, MgCl 2 , EDTA and a polysorbate-20.
10. The formulation of claim 9 , wherein HEPES is at about 40 mM.
11. The formulation of claim 9 , wherein NaCl is at about 125 mM.
12. The formulation of claim 9 , wherein KCl is at about 5 mM.
13. The formulation of claim 9 , wherein MgCl 2 is at about 5 mM.
14. The formulation of claim 9 , wherein EDTA is at about 1 mM.
15. The formulation of claim 9 , wherein polysorbate 20 is at about 0.05%.
16. The formulation of claim 9 , wherein the pH of the formulation is about 7.5.
17. The formulation of claim 9 further comprising a nucleic acid molecule.
18. The formulation of claim 17 , wherein the nucleic acid molecule is an aptamer.