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Molecular microarrays and helical peptides

a helical peptide and microarray technology, applied in the field of semiconductor lithography, can solve the problem of random conformation of peptides synthesized in vitro and on solid surfaces

Inactive Publication Date: 2010-02-11
CABEZAS EDELMIRA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This approach enables the production of biologically active microarrays with controlled three-dimensional structures, improving the biological function and activity of peptides by promoting specific conformations such as helix formation, thereby enhancing their utility in research and diagnostic applications.

Problems solved by technology

One challenge that exists in the design and implementation of peptide microarrays is the tendency of peptides synthesized in vitro and on solid surfaces to adopt random conformations.

Method used

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  • Molecular microarrays and helical peptides
  • Molecular microarrays and helical peptides
  • Molecular microarrays and helical peptides

Examples

Experimental program
Comparison scheme
Effect test

example 1

[0054]A glass substrate was silanated using a solution of 5% APTES (aminopropyl triethoxy silane) in 95% ethanol. The surface of the substrate was then washed and annealed at about 100° C. for about 1 hour. The substrate was then treated with a 5% solution of DIEA (diisopropyl ethyl amine) in DMF (dimethylformamide). A spacer molecule was then coupled by submerging the surface into a solution of O—(N-Boc-2-aminoethyl)—O′—(N-diglycolyl-2-aminoethyl) hexaethyleneglycol, 0.1 M HOBt, and 0.1 M DIC (diisopropylcarbodiimide) in NMP (N-methylpyrrolidone) with gentle agitation for about 30 min. After coupling was complete, the surface was washed with NMP. Unreacted amine groups on the surface were capped by treatment with 2% acetic anhydride in DMF solution for about 30 minutes. The surface was then washed with DMF and isopropanol.

[0055]A photoresist was prepared by mixing about 2.5% by mass of PMMA, 10% by mass of Bis(4-tert-butylphenyl)iodonium triflate and 10% by mass of isopropyl-9H-thi...

example 2

[0058]An array of wildtype (SDLHKL) and mutant (AGLHKL) peptide was synthesized on an aminated glass surface with a linker molecule, O—(N-Boc-2-aminoethyl)—O′—(N-diglycolyl-2-aminoethyl) hexaethyleneglycol, for spacing the peptides from the surface. The peptides were synthesized in a checkerboard pattern using uniform photodeprotection of t-Boc protecting groups through an open grid mask till the second leucine and spatially localized deprotection through a checkerboard mask for the last two amino acid couplings.

[0059]The photodeprotection and coupling of linker molecules and amino acids was carried out as described in Example 1.

[0060]The peptide array was incubated for 1 hour with 5 μg / ml monoclonal antibody known to specifically recognize the SDLHKL epitope of human p53 protein. A second incubation was performed with fluorescein-labeled rabbit antibody raised against mouse antibody at a 1:100 dilution in phosphate buffered saline with 0.05% Tween 20. A fluorescent checker board pa...

example 3

[0061]Photoresist formulations may include a sensitizer in addition to the photogenerated acid catalyst to generate the acid deprotection catalysts. In general, the amount of PMMA in the resist in these exemplary formulations may vary between about 3% and about 50%.

[0062]Useful photoresists may be made using diaryliodonium salts (DAI) and photosensitizers. The mass ration between DAI and photosensitizer may be between about 1:10 and 1:1. For instance, (toluylcumyl)polonium tetrakis (pentafluorophenyl) borate with isopropyl-9H-thioxanthen-9-one may be formulated in a 1:10 or 1:1 (or a ratio there between) in PMMA and PGMEA to form final concentrations of between about 0.5% to 10% by mass DAI. The formulation selected may be spun coated on the substrate surface and baked. The radiation exposure dose may be between about 0.02 J and about 10 J. Post exposure baking may be conducted for about 30 to 60 seconds at about 40° C. to about 85° C.

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Abstract

Methods for fabricating dense arrays of polymeric molecules in a highly multiplexed manner are provided using semiconductor-processing-derived lithographic methods. Advantageously, the methods are adaptable to the synthesis of a variety of polymeric compounds. For example, arrays of branched peptides and polymers joined by peptide bonds may be fabricated in a highly multiplexed manner. Additionally, peptides that adopt helical structures are synthesized on a substrate surface and arrays are created having one or more features containing peptides capable of forming helixes.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]The present application is a Divisional application of U.S. application Ser. No. 11 / 641,244 entitled “Molecular Microarrays and Helical Peptides,” filed Dec. 19, 2006 now pending. The present application is related to U.S. application Ser. No. 11 / 395,899, entitled “Massively Parallel Synthesis of Proteinaceous Biomolecules,” filed Mar. 30, 2006, now pending, which is a continuation-in-part of U.S. application Ser. No. 11 / 291,296, filed Nov. 30, 2005, entitled “Massively Parallel Synthesis of Proteinaceous Biomolecules,” filed Jun. 6, 2005, now pending; U.S. application Ser. No. 11 / 322,268, entitled “Massively Parallel Synthesis of Biopolymeric Arrays,” filed Dec. 29, 2005, now pending; U.S. application Ser. No. 11 / 529,573, entitled “Method for High Throughput, High Volume Manufacturing of Biomolecule Micro Arrays,” filed Sep. 29, 2006, now pending; and U.S. application Ser. No. 11 / 585,413, entitled “Solid-phase Mediated Synthesis of Molec...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C40B50/18
CPCG01N2001/061G01N1/06
Inventor CABEZAS, EDELMIRA
Owner CABEZAS EDELMIRA
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