Creating a Flexible Automation Platform for Plate Replication and Cell-Based Screening
M. DeGuzman
A. Bhattacharya
R. Berman
Velocity11
S. Capretta
G. Scheel
M. Pfeifer
C. Bley
Novartis Pharma AG
Abstract
At the Novartis® Basel Screening Operation Group in Discovery Technology, the number of wells processed by cell-based and biochemical assay screening is increasing at a vigorous rate. To facilitate the handling of these samples, a plate replication system (PRS) is needed to store a full copy of their current compound library with the ability to replicate portions of that library on demand for further processing. To tackle their current needs and projected future growth, Novartis® is collaborating with Velocity11® to design and build a fully automated platform to address their various requirements including the capability to grow with changing technology and assay types, the ability to run unattended with throughputs from 300,000 to over 1 million wells per day, perform cell-based screening in 1536 well plates, and the ability to interface with their in-house Laboratory Information Management System (LIMS).
This has proven to be a challenging task from both hardware and software perspectives. Velocity11® already produces a versatile automated platform with open-architecture software capable of integrating off-the-shelf peripherals. However, in order to provide a system that could perform 384 and 1536 well plate replication as well as cell based assays, several new hardware and software development projects had to be pursued. Some of these include the integration of several LiCONiC StoreX™ plate storage systems, the Labcyte® Echo 550 liquid transfer station, the transferring of information between the automated system and the Novartis IT system, the logical pooling of devices, and the management of reagents.
We will discuss these challenges and the creative ways in which we dealt with them in order to create a fully automated and flexible system.
Introduction
The Basel Screening Operation group in Discovery Technology of the Novartis Institutes for Biomedical Research has a vast compound library utilized by internal programs and groups to generate high quality hits and lead optimization information for further investigation as potential drug candidates. The group must rapidly and efficiently identify the compounds which offer the best potential to be effective lifesaving drugs. This can only be accomplished by combining cutting edge computational, automation, and screening techniques to rapidly mine through the large chemical libraries and ever-increasing number of targets to find the best match.
Maintaining this library as well as providing others with replicates and assay plates is a vital but time consuming process for the group. To help address their current needs as well as prepare for future growth, the group has approached Velocity11®.
Velocity11® is an automation company that specializes in innovative automation solutions. As well as designing, developing, and building their own automation, Velocity11® develops fully integrated automated systems based on the BioCel® 1600 platform. The BioCel® platform uses a 360° accessible robot to access an array of Velocity11® products and third party devices around its center. The BioCel® is controlled by the VWorks© software package which features a dynamic scheduler and robust error handling interfaces.
Novartis® had an extensive list of processes and specifications that currently have not been developed or demonstrated on any other automated platform. With further discussion regarding this project, it became apparent that a significant amount of hardware as well as software development needed to be done by Velocity11®. The system required the integration of new third party peripherals such as the Labcyte® Echo 550, Innovadyne Technologies’ Nanodrop™, LiCONiC Store X™ 500/1000, and the Genetix aliQuot™. The design and manufacturing of new hardware such as the Lid Hotel™ (a plate de-lidder and re-lidder), the Plate Rotation Shuttle™ (a reorientating shuttle for dual robot access), a new multi-colored Status Light System, a Cooling Jacket™, a reagent bottle cooling system, and the Weigh Pad™ (a reagent bottle liquid level sensing pad) easily address various parts of the procedure that are not dealt with by any robotics or automation to date. VWorks© needed to become highly integrated with the Novartis® LIMS to query, update, and track candidates from library to assay plate while they were being processed. Recording and tracking volumes within source plates also needed to be incorporated within the software. VWorks© error handling functionality had to be expanded, and VWorks© needed to be further developed to allow multiple devices on the BioCel® to be logically pooled so that identical devices would perform the same process on different plates.
Materials and Methods
The BioCel® is designed to be a modular system. This allows for easy access to all devices for service and maintenance, provides the ability to swap out pieces of equipment as technologies evolve, and keeps the system highly flexible. For Novartis®, the main function of the BioCel® will be to store and renew a full copy of the compound library as DMSO solutions (source plates) at regular intervals. It will also replicate on demand small aliquots of these compound solutions into destination plates which will subsequently serve as entry plates for various automated and semi-automated assay platforms. These sets are called a runset. The runsets are processed sequentially; however the system will be able to schedule future runsets while the system is busy processing a different runset.
Runset processing usually consists of compound transfers and the addition of reagents. Some examples of these reagents are salt-containing buffers, cell culture media, protein solutions, DMSO, and controls. The scheduler will determine which liquid handler will perform this function, based on the volume and specific reagent required.
Table 1 is a listing of all of the major components of the system with a diagram (fig. 1) of its placement on the system:
| Table 1: Major System Components | |||
| SUBSYSTEM | COMPONENT | QTY | FUNCTION |
| Plate Transfer | BioCel® 1600 Platform | 2 | 360° workspace with high speed robotic plate handler (A) |
| Liquid Handling | VPrep® Pipetting Station - 384 Fixed Tip Head | 4 | Low volume compound transfer (50nl - 30µl) (B) |
| Labcyte® Echo 550 | 1 | Low volume compound transfer (5nl - 50nl) (C) | |
| Reagent Distribution | Innovadyne Nanodrop© | 1 | Low volume reagent distribution (50nl - 2µl) |
| Genetix aliQuot™ | 3 | Bulk reagent distribution (2µl - 30µl) | |
| Bottle Sensing | Velocity11® Weigh Pad™ | 6 | Reagent and wash fluid monitoring |
| Centrifuge | VSpin™ | 1 | Front-loading plate centrifuge |
| Shaker | H+P Labortechnik TELESHAKE 1536™ Plate Shaker | 4 | High speed 1536 plate shaker |
| Plate Storage | LiCONiC® 1000 - Temp/Humid Ctrl. | 1 | 1000 plate, climate-controlled environment storage unit (D) |
| LiCONiC® 500 - Temp/Humid Ctrl. | 1 | 500 plate, climate-controlled environment storage unit (E) | |
| VStack™ Modular Labware Stacker | 6 | 50 plate capacity, room temperature | |
| Plate Incubator | LiCONiC® 500 - Temp/Humid/CO2 Ctrl | 1 | 500 plate, temperature & CO2 controlled storage unit (F) |
| Lid Handling | Velocity11® Lid Hotel™ | 20 | Lid removal and storage (G) |
| Other | Plate Hotel | 1 | 10 position deck-mounted plate hotels |
| Barcode Reading Station | 1 | West-side barcode reading station | |
| Figure 1: Front Isometric View of BioCell |
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The system will be comprised of different types of liquid handlers to manage a diverse volume set. The system will utilize the Echo 550™ to transfer compounds as small as 5nl and the Nanodrop™ will be capable of dispensing 50nl volumes of bulk reagents to 1536 well plates. For 384 well plates, the VPrep™ and aliQuot™ will perform similar functions on a larger volume scale. All of these liquid handlers are capable of dispensing the required volumes with a CV of less than 10%.
| Figure 2: Dynamic Range of the Liquid Handlers on the BioCel |
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[Note: The Echo 550™ and the VPrep™ perform compound transfers and the Nanodrop™ and aliQuot™ dispense bulk reagents.]
Climate-controlled plate incubator units will be capable of holding all of the plates in up to three different conditions. For example, destination plates which contain cells will need 37°C/5% CO2. Cell-free destination plates can either be stored in an incubator cooled to 4°C or in plate stackers at room temperature. With all of these storage units, the system is capable of storing and processing 2400 plates.
Assay-specific destination plates will be loaded by the user into the system’s input storage units each day. After the addition of compound solution and assay-specific reagents and controls, the processed destination plates will move to the output storage unit of the system. From there, they will be transported by the user to various assay platforms.
Several processes will be programmed on the BioCel®. The process that will be used is determined by both the number of wells and types of source and destinations. Additionally, the system will be capable of expanding and collapsing compounds between 384 and 1536 well plates.
A typical process is illustrated below:
| Figure 3: Outline of a Typical Process Performed on BioCel |
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[Note: Source and destination plates can be either 384 or 1536 well plates, which will also dictate the type of liquid handlers used.]
Results
Since the system needs to interface with many different components of varying sizes, the BioCel is actually comprised of two robotic cells. The main cell is a standard 1600 BioCel® that contains the majority of the deck components. The secondary cell is a smaller tabletop with a robotic arm to access all of the storage units and the Echo 550™. With this design, we are able to keep the footprint as small as possible while maximizing the throughput of the system.
| Figure 4. A Top-Down View of the BioCel® |
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With the advent of new peripherals as well as new techniques, several new types of hardware items were designed and developed by the engineers at Velocity11®. One of these is the Lid Hotel™ (figure 5) which was created in conjunction with Novartis®. This passive lid hotel allows the robot to remove lids from plates and then re-lid the plates when all process steps are completed. The same plate will always return to the same lid.
| Figure 5: Lid Hotel™ |
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[Note: The Lid Hotel™ is a passive lid restraint system that allows the BioCel® to de-lid and re-lid plates. Concept developed by Novartis Pharma AG®, NIBR/DT/IAT, Basel, Switzerland.]
Another new design is the Plate Rotator Shuttle™. This item allows the robot to transfer plates between the two cells while preserving orientation. Both arms can place and retrieve plates in the shuttle at the same time.
Some other requirements on the system that needed to be addressed were reagent management and reagent cooling. A new cooling jacket (figure 6) was designed to keep bulk reagents cool by using recirculating coolant while a new cooling shelf was created for the VPrep to keep source plates or reagents in reservoirs between 4 - 10°C.
| Figure 6: Cooling Jacket Designed to Keep Bulk Reagents Cool |
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To monitor the levels of these reagents, especially those under the deck, a new Weigh Pad™ (figure 7) was developed that monitors reagent bottles ranging in size from 500ml to 20L by weight. When the weigh pad senses that the liquid level is above a preset maximum or below a preset minimum level, an alarm or notice is sent to the user that it is time to empty waste or refill reagents.
| Figure 7: Weigh Pad™ Designed to Manage Reagents |
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In addition to all of the hardware development, a significant portion of the project is also software based. Order handling, process data management and process control are key requirements identified by the Novartis Information Technology group. The specifications called out new graphical user interfaces for system operation, set-up, error-recovery and the link to the different process devices but also a link to the Novartis® IT environment.
The BioCel® is operated through Velocity11’s VWorks© software platform. VWorks is the main user interface and task scheduler for all BioCel systems. This software provides a flexible base on which to integrate new devices and features to meet custom needs. As this plate replication system is positioned in a critical path within the Novartis® IT data flow, VWorks© has to be able to accept and disseminate plate, runset, and system information to and from the Novartis® LIMS (concept developed by Novartis Pharma AG®, NIBR, DT, IAT, Basel, Switzerland). This entailed the development of a plugin interface to the VWorks software in order to properly interface to the Novartis® LIMS. This integration occurs on three levels: plate (source and destination plate data), runset (data in regards to processing), and error level (any error state of the system if the system status changes).
With the integration of the LiCONiC Store X™ units, Velocity11® has created a plate database that the Novartis plugin can access to monitor the plate inventory (within the units) that was created in conjunction with Novartis®. The plate database has an Inventory User Interface which allows the end user to monitor and edit the entire plate inventory. The end user will have the ability to check the full inventory or any portion of the source plate inventory at any time. During this check, VWorks© retrieves plate specific data based on barcode from the Novartis® Database.
| Figure 8. Screen Shot of Plate Database |
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[Note: The plate database contains information about barcode, location, and liquid volume.]
Access to a plate storage system with individually addressed plate locations has added a powerful new feature to the functionality of VWorks©. By keeping an inventory of the stored plate library, VWorks© will allow the Novartis® end user to select plates by library index for use in a protocol.As source plates pass through the system, the VWorks© inventory functionality maintains an accurate record of the volume remaining in each quadrant of the source plate. VWorks’ added volume tracking functionality (concept developed by Novartis Pharma AG®, NIBR, DT, IAT, Basel, Switzerland) allows the Novartis® end user to program a low volume threshold. When a source plate falls below this threshold, an error flag is generated for that plate within the inventory.
A new software feature also designed for this system allows the user to select from a device pool of instruments with multiple instances on the system. This feature is best displayed with the BioCel’s VPrep™s. The user can specify any of the four VPrep™s in the system to simultaneously run the same pipetting protocol in order to raise the system’s throughput and provide redundancy. VWorks© will pool the VPreps specified and use the next available VPrep as it continues through the process. This device pooling feature will also allow for multiple instances of an instrument to act as a backup for other instruments. In the case of VPrep™s, if one VPrep encounters an error, VWorks© will report the error and stop the process for that VPrep™ only. However, the VWorks© scheduler will continue to process plates on all other VPrep™s specified for that particular pipetting task continuing the work flow.
Discussion and Conclusion
The BioCel® has already proven itself to be a versatile platform with the ability to adopt new technologies and expand current software and scheduling capabilities to meet any customer's needs. The engineers at Velocity11® were able to create new hardware to add to the functionality and robustness of the system. Working closely with Novartis® and understanding their needs has enabled Velocity11® to provide them with one of the most innovative automation solutions available. This advancement will enable Novartis® to streamline their procedure for source plate replication in preparation for high throughput screening.
With the extensive hardware and software development accomplished by this project, Novartis' process has become more automated and integrated with their in house LIMS system. This will insure the system is integrated with processes, databases, and software already in place. Moreover, the custom system will give them significant productivity advantages as they will be able to process plates up to 1536 wells using a variety of assays. Additionally, since the system will be capable of running unattended overnight, Novartis will be able to process up to 800 plates in 24 hours or over 1 million compounds per day.
Acknowledgements
We would like to acknowledge the tremendous efforts of Velocity11’s software and custom engineering teams. Contact Information:
Maria DeGuzman, Velocity11®
mdeguzman@velocity11.com
