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Producing high-density peptide arrays

Feb 2008
Gary Boas

High-density peptide arrays could be used in a wide variety of applications. Improving the state of the art from the existing 22 peptides per square centimeter has proved challenging, however. Researchers typically create the arrays by coupling monomers to the solid support consecutively. Generating arrays from the 20 different amino acid monomers therefore involves an inordinate number of coupling cycles, adding to the overall complexity of the process.

To reduce the number of cycles, and thus clear a path for high-density peptide arrays, a team with the German Cancer Research Center and with the University of Heidelberg, both in Heidelberg, developed an alternative means of creating the arrays.


Researchers have reported a method that not only reduces the number of coupling cycles to generate a peptide array but also the overall complexity of the process. Rather than coupling monomers to the surface consecutively, the chemical reaction is “frozen” inside particles (A); coupling occurs only after an entire layer of particles has been moved to the surface (B) and has melted (C). An array is created through multiple cycles of this process (D). Images reprinted with permission of Science.

The researchers produced 20 kinds of chargeable amino acid particles, which were guided, step by step, onto the microchip surface by electric field patterns from individual pixel electrodes. The chemical reaction essentially is frozen inside the particles, and coupling occurs only after a layer of particles has been moved to the surface and melted. “This principle allows for the miniaturization of the whole process and at the same time reduces the number of coupling cycles to one per cycle,” said Frank Breitling, who, along with Ralf Bischoff and Volker Stadler, is a principal investigator.

Breitling described the challenge of developing the particles: The researchers needed to produce particles that are chargeable and to find ways to maintain the charge over time while not interfering with the chemistry of peptide synthesis. He and his co-workers addressed this challenge first by studying toner particles from a commercial laser printer “to learn about the physical properties our amino acid particles should have.” Then they tested various ingredients used to manufacture such particles for interference with the peptide synthesis. They produced a formulation that had the physical properties of commercial laser printer toner but that did not interfere with the coupling reactions.

The investigators compared the new method with the conventional Merrifield synthesis and observed similar yields of synthesized proteins, with negligible decay rates for 19 of the 20 amino acid particles. Then they synthesized an array of peptides differently labeled with FLAG- and HA-specific antibodies on the surface of a microchip, producing an epitope-specific staining pattern with a density of 40,000 peptide spots per square centimeter.

Using the method, the researchers synthesized an array of peptides labeled with FLAG- (green) and HA-specific (red) antibodies with a density of 40,000 peptide spots per square centimeter.

The technique could advance a wide variety of clinical and research applications. Breitling noted, for example, that investigators could use it to synthesize a large conformational space -- that is, to synthesize a large number of different peptides on an array, all with varying three-dimensional conformations, and then to screen for binders. “The more peptides we have on the array, the likelier we are to find [good] binders,” he explained.

The researchers continue to develop the method. They plan to produce a small instrument with a chip-print head that can generate high-density peptide arrays inexpensively. They also are working to miniaturize synthesis sites further, with a final goal of 1 million peptides per square centimeter.

Science, Dec. 21, 2007, p. 1888.

amino acid particlesBasic ScienceBiophotonicsHigh-density peptide arraysmonomersNews & Features

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