Substrates, systems, and methods for array synthesis and biomolecular analysis
Disclosed herein are formulations, substrates, and arrays. In certain embodiments, substrates and arrays comprise a porous layer for synthesis and attachment of polymers or biomolecules. Also disclosed herein are methods for manufacturing and using the formulations, substrates, and arrays, including porous arrays. Also disclosed herein are formulations and methods for one-step coupling, e.g., for synthesis of peptides in an N->C orientation. In some embodiments, disclosed herein are formulations and methods for high efficiency coupling of biomolecules to a substrate.
1. A method of using a photobase generator for attaching a coupling molecule on a surface of an array, comprising:
obtaining a substrate of an array comprising a plurality of carboxylic acid groups for linking to a coupling molecule;
contacting the substrate with a carboxylic acid activating compound formulation;
contacting the carboxylic acid activated substrate with a photoactive coupling formulation;
selectively exposing the photoactive coupling formulation to radiation; and
coupling the coupling molecule to at least one of the plurality of carboxylic acid groups at the selectively exposed area,
wherein the photoactive coupling formulation comprises:
(a) a photobase generator comprising a compound of formula (II):
wherein
is a nitrogen-containing cation comprising
X is NH or CH 2 ,
n is 0 to 3,
R 5 is aryl or heteroaryl,
A ⊖ is
or tetraphenylborate,
R 6 is hydrogen or —NO 2 ; and
(b) the coupling molecule comprising an amino group that is coupled to a protecting group;
and wherein upon exposure of said photoactive coupling formulation to radiation, said protecting group is removed from said coupling molecule, and said unprotected coupling molecule binds to at least one of the plurality of carboxylic acid groups that are attached to the carboxylic acid activated substrate.
2. The method of claim 1 , wherein said anion A ⊖ is tetraphenylborate.
3. The method of claim 1 , wherein said anion A ⊖ is a phenylglyoxylate.
4. The method of claim 1 , wherein said compound of formula (II) is selected from the group consisting of compounds of the formula:
5. The method of claim 1 , wherein the photoactive coupling formulation comprises a polymer and the coupling molecule is an amino acid.
6. The method of claim 5 , wherein said amino acid comprises said protecting group.
7. The method of claim 6 , wherein said protecting group is base labile.
8. The method of claim 6 , wherein said protecting group is Fmoc.
9. The method of claim 5 , wherein said amino acid is present at 0.1M in said photobase composition.
10. The method of claim 5 , wherein said polymer is present at 0.5-3% in said photobase composition.
11. The method of claim 5 , wherein said polymer is polymethyl methacrylate.
12. The method of claim 1 , further comprising repeating said method to produce a desired polymer at said at least one carboxylic acid group.
13. The method of claim 1 , wherein said coupling step is performed multiple times at different selectively exposed areas on said substrate.
14. The method of claim 1 , wherein said coupling step has a coupling efficiency of at least 98.5%.
15. The method of claim 1 , wherein said substrate comprises a planar layer comprising a metal and having an upper surface and a lower surface; and a plurality of pillars operatively coupled to the layer in positionally-defined locations, wherein each pillar has a planar surface extended from the layer, wherein the distance between the surface of each pillar and the upper surface of the layer is between 1,000-5,000 angstroms, wherein the surface of each pillar is parallel to the upper surface of the layer, and wherein the plurality of pillars are present at a density of greater than 10,000/cm 2 , and wherein the attachment site is coupled to the upper surface of the pillar.