Abstract
With drug discovery technology advancing and changing rapidly, there is a growing need for automated equipment to have the capability to change with these changes in technology. The use of single workstation approaches to liquid handling is decreasing due to the increasing need for a system that can perform various liquid-handling tasks on the same platform.
The Staccato™ Automated Workstation is an extremely versatile liquid handler with the capabilities of handling plate formats ranging from 96 well to 1536 well plates. The ‘Drag and Drop’ architecture of the software allows for flexibility and ease in process development. With the use of CLARA™ 2001 software, method development and system reconfiguration becomes less intensive. Its scalability lends itself to supporting changes in platforms as emerging technologies arise.
The Staccato™ system can be equipped or configured with many components and options to fit liquid handling needs. Using the SciClone ALH as the base liquid handler, the system can come equipped with a disposable tip Rapid head, or a small volume fixed tip head with 96 or 384 cannula arrays. To augment the liquid handling capabilities of the SciClone, up to four separate Cavro™ bulk dispensing manifolds can be mounted on the head for reagent additions. Liquid handling can be further enhanced by the addition of an eight tip, independent Z-axis probe used for various liquid handling applications. Integration of the Autostack with the SciClone provides exceptionally large storage capacity for unattended operation. The use of on-board barcode readers in conjunction with the CLARA™ Data Manager, gives the system the ability to track all compound and assay plates entering the system.
Reformatting chemical libraries with the Staccato™ system is very efficient. With its large storage capacity and flexible configurations, reformatting can be done through a variety of methods. Using the Rapid head configuration, 300 96 well plates can be reformatted to 75 384 well plates in a single eight hour day. Using the small volume fixed tip configuration, one can convert 200 96 well plates to 50 384 well plates in a single eight hour day. Although fixed tip configurations yield less throughput due to implementation of wash protocols, there are no consumable costs. It also provides the ability to further reformat from 384 to 1536 well plates, with an even smaller volume 384 cannula array designed for 1536 applications.
The independent Z-axis probes provide the ability to cherry pick select compounds, assemble serial dilution plates, and manipulate compounds and reagents for ADMET studies. Performing parallel processing of plate types using the ‘on the fly’ array swap capabilities, the Staccato™ is capable of reformatting from 96 to 384 well plates using the 96 cannula array. After dropping off the 96 array, it can then pick up the 384 array and further dilute, or deliver compounds directly to assay plates for execution with no human intervention.
The presented pipetting specifications are based on five trays run with three transfers performed per tray. A 0.5μl wet transfer produces excellent precision with an average percentage CV of 6.4, and an accuracy range of 1.06 to 7.3%. A 0.5μl dry transfer yields an average precision percentage CV of 8.6, and an accuracy range of 0.03 to 10.1%. 1μl dry transfers yield a precision average percentage CV of 5.8 and an accuracy range of −1.5 to 4.4%. Volumes larger than 1μl yield similar results to the 1μl numbers.
System use at Neurogen Corporation entails taking in chemical entities from our existing archive or from our high-speed synthesis group and performing compound dilution procedures, or reformatting processes on the plates and transferring the compounds directly into assay plates, all within the same method. On this platform, the system can also build dose response plates, prepare compounds for inter-departmental support, or execute a variety of assays.
After compounds are in the proper format and concentration, with the use of the Cavro™ dispensers, multiple reagent addition to assay plates can be accomplished with the proper scheduling of processes. In essence, the system can take compounds from their starting format and concentrations, manipulate the plates to accommodate assay specific plate formats and conditions, then proceed to add the appropriate reagents via the Cavro™ dispensers, bringing the assay from compound delivery, through reagent addition. Dispensing by the Cavro™ syringes for volumes of 10μl-2500μl all yield similar results. Across plate precision yields an average percentage CV of 3.8 with an accuracy range of 1.1 to 5.5%. Non-contact dispensing of reagents eliminates cross well contamination and with multiple manifolds, various reagents can be used.
Replication of compound data generated by the 384 cannula array at a dry transfer volume of 0.5μl is quite good. Dry dispensing of such a small volume is not easily achieved, but obtainable with acceptable percentage CV using particular pipetting methods. Using the same pipetting methods and comparing data to proven control methods for drug delivery, yields very acceptable percentage CV's and correlation.
CLARA™ 2001 Software is used for scheduling methods and integrating peripherals into the existing system. CLARA™ communicates with adapter modules that translate commands between CLARA™ and the ICP of the peripheral module, triggering program execution of the instrument. This method of integration makes the scalability of the system much easier and more efficient than previous, conventional methods. Upon process completion, the ICP sends a message back to CLARA™ through the adapter to confirm completion, or send notice of an error in execution. The system's open architecture, in conjunction with the CLARA™ software, allows for flexibility and ease when modifying or implementing new assay platforms and technologies.
The CLARA™ Data Manager provides the ability to manipulate and filter incoming data with the use of the Data Manager's ZyMap. Data can be configured into text files, excel sheets, or directly transported to a database via ODBC connections. However, the current version of the Data Manager is unable to catch critical events that would trigger the data to be sent to the database, such as an ‘end of run’ event. This inability does not yet allow the direct triggers to export data to a database. A temporary remedy supplied by Zymark is a developer's kit (a collection of APIs) that gives you the ability to build your own Data Manager using pre-defined templates. Using this method could require extensive time and programming on the user's end depending on the process and data being captured. However, an updated version of the Data Manager is anticipated to be released at the end of October that would alleviate this problem.
Although the Staccato™ system has many benefits and capabilities for liquid handling needs, there were some implementation issues along the way. The biggest setback experienced was with limitations of the Data Manager. One of the top reasons for the selection of the Zymark system over other vendors’ was the ease of database connectivity and data export. Another issue was in the design of the fixed tip arrays. Great care must be taken not to damage their Teflon coating. Any scarring or scraping of the material will cause variation in pipetting procedures especially at small volumes. Service for the most part is good; however, the redesign and complexity of the system left the need for more time in training of technicians to come up to speed in the overall workings of the system.
Overall Neurogens experience with the Staccato™ system has been a good one. All systems have their benefits and drawbacks. The most important thing is the cooperation and speed of assistance received when problems do arise and efficiency in coming up with solutions. In this aspect, Zymark has shown dedication and commitment to their products and customers.
Acknowledgements
I would like to thank the scientists who helped in validating the system: Richard Yanicky for all his help with data capture and export, and Dave Scanlon along with Zymark technical support for all their help along the way.