DOC-P

The Digital Optical Chemistry-Proteomics (DOC-P) System is designed to rapidly manufacture high-density protein detecting arrays (PDA). DOC-P is an expansion of the proven Digital Optical Chemistry (DOC) System used to build oligo arrays for detecting DNA sequences. For work with proteins, peptoid capture ligands are assembled on a standard glass slide in a programmed sequence using ultraviolet photochemistry. Protein samples are then hybridized onto the protein-detecting array and labeled with a fluorescent marker. The labeled array is scanned using standard equipment and a sandwich type assay is performed.

Protein Capture and Labeling Sequence

The DOC-P is currently configured to deliver twenty-four different chemicals to be used in the synthesis of peptoid arrays. The chemicals are delivered to the reaction chamber via computer controlled valves. There is expected to be twenty positions available for amino acids as some of the chemicals will be solvents and reagents needed to facilitate the reactions. There are eighteen additional control lines available that may be used if additional chemical selections are desired; however, valves and plumbing would have to be added.
The DOC-P reaction chamber is mounted in a conventional 500 watt microwave oven that operates at 2.45Ghz. The duty cycle of the oven is adjustable to allow the power delivered to the reaction to be varied if necessary. The microwave stimulated reactions are expected to require minutes to reach completion versus hours without microwave stimulation.
Peptoid arrays are built by sequentially adding amino acids at sites in the array determined by an ultraviolet image projected onto the array. The amino acid for addition is introduced into the reaction chamber under program control. The image corresponding to the sequence in the array is projected onto the slide in the reaction chamber and sites in the array that are illuminated will have the amino acid added to their sequence. The cycle of amino acid addition repeats until the peptoid probe array is complete. The image or mask series used to expose the array is generated by a separate probe design software package and are supplied to the DOC-P operating program each time a different array is to be built.

DOC-P Functional Diagram

The mask image is projected using a Texas Instruments Digital Light Processor (DLP). The DLP is a 768 by 1024 array of mirrors that are controlled by an image supplied by the computer. A collimated beam of light is directed onto the DLP and the individual mirrors will reflect the light either into the reaction chamber when selected or off to the side when deselected. Each mirror is 13 microns square with a total of 786,432 mirrors in an area of approximately 12mm by 15mm. Currently, the DOC System which uses the same DLP imaging scheme has been used to build arrays of approximately 200,000 oligos with 2 by 2 mirror patterns (25 by 25 microns). The potential exists to increase the array density with some tuning of the optical system and chemical processes.
The ultraviolet light is supplied by a 1000 watt mercury vapor arc lamp. The output of the lamp is filtered to pass only the 365 nanometer line that is used for the photochemistry. The filtered light is integrated then focused onto the DLP.

DOC-P Imaging System

The DOC-P is fully automated allowing the operator to start the program and return to retrieve the completed peptoid probe array. The operation is expected to take approximately five minutes per amino acid addition. With additional time required for supporting functions such as washing and drying the reaction chamber, an array is expected to be completed in a few hours, depending on the length of the peptoid probe sequence.
To build a protein detecting array and analyze protein samples requires supporting functions at the front and back end of the DOC-P System peptoid probe array fabrication. The following sequence is representative of the procedure required to perform a complete analysis of a protein sample using a peptoid probe array:

Flow Diagram for Protein Analysis Using Peptoid Arrays

The DOC-P System hardware is complete and the basic functionality has been tested. The following testing and development in support of the DOC-P System has been completed to enable preliminary peptoid array fabrication and analysis:

• The fluidics system has been tested under conditions that are expected when running the peptoid chemistry.
• The optical system has been aligned and focused using the existing DNA chemistry (see focus image).
• The microwave functionality and power levels have been tested.
• A Preliminary DOC-P operating system has been developed.
• Preliminary probe design software has been developed to generate the masks and sequence listing needed to run the DOC-P and analyze the arrays.

Work now centers on testing various operating scenarios in anticipation of the peptoid chemistry and ligand synthesis. Currently, operating conditions simulating the amino acid addition cycles are being evaluated for temperature changes at the reaction chamber.

DOC-P Focus Test Pattern Using DNA Chemistry – Each Square is 90 X 90 microns

Glimpse of a DOC-P Work Station