Oxide layers on silicon substrates for effective confocal laser microscopy
View Patent ↗Methods of performing confocal laser microscopy on a polymer array disposed on a silicon wafer substrate, the method comprising the steps of providing a silicon wafer substrate having a top side and a bottom side, coating the top side of the silicon wafer with an oxide coating to provide an oxide coated wafer, covalently coupling a plurality of probes to the top side of the coated wafer to provide a fixed polymer array, hybridizing the fixed polymer array with a plurality of labeled ligands, and assaying for one or more hybridized ligands using confocal laser fluorescence microscopy to detect hybridization are provided.
1. A method of performing confocal laser microscopy on a polymer array disposed on a silicon wafer substrate, the method comprising the steps of:
providing a silicon wafer substrate having a top side and a bottom side;
coating the top side of the substrate with a transparent oxide layer to provide an oxide coated wafer;
depositing a reactive functional group comprising a labile protecting group substantially uniformly across the transparent oxide layer, wherein the labile protecting group is NNPOC or MBPMOC;
selectively removing one or more of the labile protecting groups from predefined regions of the wafer to provide exposed functional groups in the predefined regions;
reacting the exposed functional groups with a nucleic acid comprising a reactive functional group and a labile protecting group;
repeating the steps of selectively removing and reacting to produce the polymer array;
hybridizing the polymer array with a plurality of ligands; and
assaying for one or more hybridized ligands using confocal laser fluorescence microscopy to detect hybridization.
2. The method of claim 1 , wherein the oxide layer has a thickness of at least 3,500 angstroms.
3. The method of claim 1 , wherein the oxide layer has a thickness of at least 35,000 angstroms.
4. The method of claim 1 , wherein the labile protecting group is removed by activating a photoacid generator with light of an appropriate wavelength to produce acid.
5. The method of claim 4 , wherein the photoacid generator is an ionic photoacid generator or a non-ionic photoacid generator.
6. The method of claim 4 , wherein the photoacid generator is an ionic photoacid generator.
7. The method of claim 4 , wherein the photoacid generator is a non-ionic photoacid generator.
8. The method of claim 7 , wherein the non-ionic photoacid generator is 2,6-dinitrobenzyl tosylate.
9. The method of claim 6 , wherein the ionic photoacid generator is an onium salt.
10. The method of claim 9 , wherein the onium salt is bis-(4-t-butyl phenyl) iodonium PF6 − .