Purpose:

         The AECOM Microarray Printer is a special purpose instrument designed to deposit high-density, gridded arrays of cDNA, genomic DNA or similar biological material on microscope slides. The principal components are a computer controlled high precision, high speed, three-axis robot and a unique pen tip assembly. It is capable of depositing thousands of extremely small droplets of genetic material on a small surface area, one droplet at a time, with each droplet containing a different gene. Such a capability permits gene expression experiments using tiny samples of genetic material while obtaining simultaneous data on thousands of genes.
 
 
 
 
 
 

Hardware Description:

Mechanical Slides

        The microarray printer consists of a three-axis robot mounted on a Newport Vibration Isolation Workstation table with a research-grade laser tabletop measuring three feet by five feet by two inches. This table provides a very sturdy support structure with good vibration damping properties. The tabletop is flat to within 0.004 inches and contains a uniform grid of holes at 1 inch centers, which imparts a high degree of modularity and reconfigurability to the design. The frame provides a self-supporting structure for the entire apparatus while maintaining isolation between ancillary equipment and the tabletop. Additional components can be added to provide support for an enclosure above the table should this prove necessary. The entire assembly is precisely leveled, while the frame structure below the table is utilized to hold the electronics control panel.
 

        The three-axis mechanical slides are from THK and are selected to provide the best combination possible of precision and speed consistent with the design objectives.

The X-axis slide (GL20) runs the full length of the table and is bolted to one edge of the tabletop. It features dual square rails with a precision ground 20mm pitch, 1inch diameter
lead screw capable of supporting speeds of over 1.2meters/sec while providing a position repeatability tolerance of 10 microns. The stiffness of the structure is enhanced by the use of a robust follower rail on the opposite side of the table, parallel to the driving slide. The laser table provides added stiffness and assures that the two slides maintain straightness and parallelism with minimal stresses and positional errors.
 
The Y-axis mechanical slide (KR46) is mounted perpendicular to and on top of the two X-axis slides. It rests on top of two supporting blocks, and is approximately 2 feet long. It is proportionately smaller than the X-axis slide, but, maintains a high degree of stiffness due to its all steel construction. The Y-axis lead screw is also 20mm pitch and is capable of 1 meters/sec speed while providing a position repeatability tolerance of 3 microns.

The Z-axis mechanical slide consists of a high precision miniature slide (KR20) mounted perpendicular to both the X-axis and Y-axis using a third mounting block resting on the Y-axis slide. It provides 5 inches of vertical travel and features a 1mm pitch, precision lead screw while providing a position repeatability tolerance of 3 microns.

Pen Tip Assembly:
 

The Pen Tip Assembly consists of a proprietary design having 12 pen tips contained within a precision-machined housing that is supported by the moving block on the Z-axis slide. The housing can be aligned precisely to insure that the pen tips move within the housing only in the direction perpendicular to the surface. Key features include: the ability to precisely hold the relative position in the X and Y direction while allowing a high degree of freedom of motion in the vertical  direction, high strength and low-mass spring-loaded pen tips that can be individually aligned precisely in the vertical direction, and controlled geometry pen tips that pick up precise quantities of material and deposit uniform, repeatable spots. The assembly can work with any number of pens up to 12, but practical usable numbers are limited to 1, 2, 3, 4, 6, 8 and 12 due to titer plate geometry. An important design goal is to be able to fabricate low wear pen tips that deposit uniform dot sizes, deposit at least 100 drops between refills while minimizing excessive waste of material.
 
Electronics:

The mechanical slides are positioned with motors and servo amplifiers from Yaskawa. These consist of a 1 horsepower (750Watt) motor for the X-axis with a rated speed of 3000RPM and 338oz-in torque; a 200Watt motor for the Y-axis rated at 3000 RPM and 90 oz-in torque and a 100Watt motor for the Z-axis with integral brake, rated at 3000 RPM and 45 oz-in torque. All three motors incorporate a 2048 count relative encoder which, through 4X phase detection, can yield an electrical resolution of 8192 counts per motor revolution. This corresponds to a linear resolution of +/- 2.5 microns for both the X-axis and Y-axis, and +/- 0.125 microns for the Z-axis.
 

The amplifiers are matched to the motor sizes and include built-in microprocessor control, solid state high voltage power circuits and a plug-in controller for monitoring and tuning each amplifier. The primary power is 220V 60Hz single phase provided from a 20 Amp mains with a wall mounted manual circuit breaker. Individual fuses for each line are incorporated into the control panel mounted underneath the table. The amplifiers operate in torque mode whereby the servo controller card in the PC provides +/- 10 volt analog control signals proportional to the required torque. The encoder signals are fed to both the amplifiers and the servo controller card, so that the position control loop for each axis is closed within the PC-based servo controller card.

Relays on the control panel allow the removal of power via an Emergency switch located in front of the table that can be activated by pushing the large red knob. Additional relays are included in the control panel for controlling the Z-axis brake, the drying station, and for providing a failsafe watchdog timer to remove power in case of a computer failure. A 24volt DC power supply on the control panel is used to provide the excitation for the relay coils as well as the opto-coupled status and control signals sent between the amplifiers and the PC-mounted servo controller.
 
Servo Controller:

The PC based servo controller is a four axes controller card supplied by Delta Tau. It can be programmed to execute complex servo algorithms and updates each axis at a 2.26KHz rate. Card configuration requires the proper setting of almost 100 jumpers to select the appropriate parameters for each application. In addition, there are over 500 software parameters that are set during the alignment and calibration phase that allow one to tailor the servo performance for each axis. The card can run motion control programs and multiple PLC programs (for housekeeping and asynchronous control functions) simultaneously. Opto-coupled inputs are provided to monitor the Hall effect sensors used to indicate the positive and negative limits as well as the home position for each axis. The servo controller can control the Enable function for each amplifier directly, allowing it to instantly disable the servos if a limit condition or fault is detected. The status of the various relays is fed back to the servo controller to help monitor fault conditions and aid troubleshooting. Power for the drying station is relay controlled via an output from the servo controller. Additional inputs and outputs are available if required.

Computer:

The computer is a Gateway 2000 model P5-166, a 166Mhz Pentium PC with 32Mbyte RAM, 3.2GB hard drive and 17 inch monitor. To insure reliable operation the computer is equipped with the a 350VA UPS to provide back up power for a controlled shutdown sequence in case of power loss.
 
 

The computer runs the Windows NT operating system. The control program for the robot was developed internally using the Microsoft Visual Basic Professional development package. The control program provides the user friendly front end (see photos) whereby the user enters the key coordinates of the titer plates, microscope slides and wash/dry station as well as the desired spot density, number of spots desired and number of slides to be processed, and the program selects the appropriate routines to run.

The Servo Controller card runs its own program language and was programmed separately for the appropriate motion routines. It communicates with the host computer via the ISA bus. An OCX package from Delta Tau allows the Visual Basic control program running in Windows NT to communicate with the card. The control program passes the appropriate parameters to the card, commands the running of the appropriate motion and PLC routines, and retrieves system status and motor parameters for control feedback and user display.

A DOS based program,  provided by Delta Tau,PMAC Executive, was used to set up the various card parameters and to tune the servos for the exact combination of hardware components and performance requirements for this system.

 

Table Configuration:

The tabletop was designed to afford the greatest possible flexibility in configurations. The usable work area is a rectangle that is 23 inches wide by 51 inches long, as determined by the range of travel of the mechanical slides.

The titer plate holder consists of a modular assembly that simply snaps into position with spring loaded keying pins designed to fit into the 1 inch holes on the laser table.
It can be positioned anywhere in the active area of the table in 1 inch increments. The user simply enters into the computer the x-y coordinate of the 1 inch area where well number A1 on the titer plate is located and its position is defined within a few thousands of an inch. Any number of plate holders can be used in whatever pattern is convenient for the user.

The microscope slides can also be placed anywhere on the work surface. The user aligns the edges of the 1 by 3 inch glass slides to the centers of the hole pattern on the tabletop. These can then be taped down at the two ends to immobilize them during the spotting process. The x-y coordinates of the 1 inch square area under the left side of each slide is used to identify its location for the computer. A group of contiguous slides can be mounted in a column and its location defined by entering the location of the first slide and the number of slides in the group. The wash & dry stations utilize the same titer plate holders described above to position wash trays and a drying station. These can also be located in any convenient location on the table and their coordinates entered in the computer. The wash trays typically consist of 1 each of distilled water and alcohol trays to rinse the pen tips, although this can be customized as well. The drying station can be either a fan assembly or a vacuum source that can be used to promote evaporation of remaining liquids on the pen tips.
 
 Performance:

The microarray printer is capable of rapidly moving to any x-y location on the table within 1 second. Its x-y position can be repeated to an accuracy of +/-11 microns, for any type of move. The robot can spot 12 points every 1/2 second, or, 100 slides with 12 spots each in under 60 seconds. At this rate it is possible to create 100 slides with 6400 clones each in under 11 hours, or, 100 slides with 25,000 clones each in about 42 hours.