Rise of automation

Automation is playing and ever increasing role in laboratories. Here, we find out how the Cell Culture Platform from RTS is allowing tissue culture to go robotic

The Cell Culture Platform automates the tissue culture process; a process that is vital to the development of immortalised cell lines. This new system has the RTS Assay-Platform among its antecedents, drawing as it does on a modular format driven by Sprint scheduling software. Like the Assay-Platform it has best in class instruments inside a Class II biological containment enclosure, served by a Tecan Evo 200/8 with a Robotic Manipulator arm (RoMa), a Liquid Handling Arm (LiHa) and a TeMo 3x5 96 tip plate to plate transfer module. 

The system features dynamic scheduling, courtesy of Sprint software. It has also been designed to be flexible, easy to expand and able to manage higher throughputs and even to ultimately become a large scale, flask cell culturing system. RTS say that there are numerous applications for a system with the broad capabilities of the platform, including drug transporter assays, cell transfection and cell line expansion.

A single operator working a standard working week is all that is required to run the system, as, with careful scheduling, the plates can emerge from the system at the beginning of the working day, so that for much of the time, the system operates unattended. The user can configure the platform to send its operator a warning an hour before the plates complete the assay. The software makes the process simple to use by handling the complex scheduling activities and plate tracking activities.

Bristol University commissioned a forerunner of the platform three years ago primarily for its work on The Avon Longitudinal Study of Parents and Children (ALSPAC). Since then further refinements have been made to the system to enables small-scale cell culture in micro titre plates. Following the success of its first system, Bristol University installed a second system, this time for the 1958 British Birth Cohort Study, which is managed by St. Georges Hospital, London and The Institute of Child Health, with the growth of cell lines being outsourced to the Bristol University team.

Each cell line starts off life in a single well of a 24-well plate. Bristol’s platform variant takes those cells and expands them across the rest of the plate and ultimately over several 6-well plates, by which time there will be enough cells to be able to extract an initial sample of DNA and to cryopreserve a stock sample of cells for the future.

For the first configuration, the platform comprises a Tecan EVO 200/8 with an eight fixed tip LiHa, a RoMa for handling the plates' transfers, an AbGene APS 300 thermal plate sealer and a Tecan Sapphire II plate reader inside a BigNeat class II type enclosure. Outside it, there is a Kendro Cytomat 6001 incubator, which controls temperature, humidity and CO2 levels, a six-way reagent valve and a Binder refrigerator unit to cool the growth media.

Automated tissue culture, like the cell culture platform ar Bristol University can increase cell line production

The first drug transporter application seeds and feeds three cell lines, though the system can cope with any number, limited only by incubator capacity. It will support feeding of a range of cell lines typically with culturing periods from five to 21 days. The platform is able to automate the seeding of trans-well plates with cells from a cell suspension, the re-feeding of cells in trans-well plates at user-definable time points and the transport assay itself.

The process is divided into a series of software schedules:
1. Seed and feed
2. Automated system check
3. Transport assay
4. Standard curve

Seed and feed

In this phase, the three different cell line suspensions in conical bottom tubes are transferred (seeded) into trans-well plates and incubated, with repeated automated feeding steps as defined by the user. Once the operator starts the processing, the system controls all the processes from this point. Between feeding cycles, system lines can be cleaned and sterilised by switching appropriate solutions into the system lines using the 6-way valve.  Multiple media may be used if required.

A second set of plates can be launched during this phase. The system is able to keep track of each plate, as well as the assay to which it is linked. RTS calls this the dynamic batch facility and this “chain of custody” relies on barcoding to provide clear provenance and data integrity.

When the culturing process is nearing completion, the system can be programmed to alert the operator by email or SMS. This gives the operator the chance to load the system with reagents and labware for the next phase.

Automated system check

Prior to starting the main assay, the system can perform an automated self-check to ensure that the cells and reagents are not wasted. Fluorescence plates are placed in the incubator and dye solution placed on the Evo deck by the operator. The function of the liquid handler and reader is checked by dispensing a serial dilution of a suitable dye (such as Texas Red or Lucifer Yellow) into a 96-well plate and reading the fluorescence on the integrated reader. If the calculated %CV is within acceptable limits the transport assay is automatically started. If the system fails this test, perhaps because it has run out of washer fluid or there is a blocked tip, the user is alerted so the problem can be dealt with.

Transport assay

Compounds and dye are dispensed to the top and bottom sections and incubated. Then the resulting solutions are copied to two fluorescence plates (top/bottom) per trans-well plate for cell membrane integrity tests and pooled to two injection plates per 3 trans-well plates, one from the top and another from the bottom before being subjected to mass spectrometry. The platform is able to pool these samples with Sprint handling the data and providing the operator with an output file showing the whereabouts of each sample. Once the fluorescence plates have been read by the Sapphire II, they are discarded. The injection plates are sealed and stored in a hotel at the front of the reader.

Standard curve

Standard solutions, quality control and blanks are dispensed to appropriate wells. The plate is sealed and placed in the output position for easy access by the operator. The plate is removed for subsequent analysis by LC/MS.

By Mel Whiteside, applications consultant, RTS Life Science.  Mel has been an Assay-Platform project manager at RTS for over three years.