Automation tames—or at least tempers—some of the many challenges of working with cell-based assays. From cell preparation and processing to assay protocols, many steps can be automated. The areas to automate that will provide the biggest return depend on the application, and cell-based assays include a wide range of uses in basic life sciences and medical research, including drug discovery and development.
When asked about the main applications of cell-based assays, Eddie Wehri—scientist, high-throughput screening at the Center for Emerging and Neglected Diseases —says, “They are endless and depend on your field but the more common would likely be potency assay for determining biological activity, mechanism of action, gene reporter, biomarker, cytotoxicity, and cell proliferation.”
Also, cell-based assays can be run in various ways, including high-throughput and high-content screening (HTS and HCS, respectively). The cells themselves also vary. They can be alive or fixed, adhering to a surface or floating in a suspension. The method of analysis varies, as well. “What are you measuring?” Wehri asked. “Is it just a simple fluorescence intensity readout, which can be done on a plate reader, or is it a phenotypic screen, like a translocation assay, which requires imaging and analysis software.”
With so many different ways of going about cell-based assays, scientists also use a broad collection of tools and technologies. In automating these assays, scientists use robotics and scheduling software. Various forms of imaging might also be involved. Plus, the assay technology itself must be considered.
As explained by Rémi Magnan, associate director cellomics and proteomics at Tecan, “The large variety of assay types makes it a challenge to provide a unique automation solution that would fit all cell based–assay formats. Depending on the final combination of assay, cell type, and readout, the design of an automated workstation can go from fairly basic to extremely complex, depending on the level of integration required.” He adds, “Tecan offers both off-the shelf as well as fully customized solutions to automate cell-based assays.”
Hurdles to handle
The challenges in automating cell-based assays depend on what is being done, and scientists keep trying more approaches. One exciting area involves assays on single cells. Despite the great potential benefits of this concept, it’s not easy to do. “The key challenges with single-cell assays involve ensuring that approximately an equal volume of cells is dispensed per well,” says Fakhar Singhera, HTS robotics engineer at Scripps Research, Florida. “This can be complicated by cells that are denser than media and thus settle down near the dispense nozzles, leading to clogging in the nozzle tips.”
Image: A range of robotic processes can be used to automate various steps in cell-based assays. Image courtesy of Fakhar Singhera.
Keeping cells in dispense bottles can also create viability problems. “As the cells aren’t generally in a temperature and humidified environment while waiting to be dispensed, cell death is a limiting factor on batch sizes for high-throughput screening and can dictate whether 20, 30, 60, or more plates can be dispensed at a given time,” Singhera adds.
To deal with these dispense-bottle issues, Singhera and his colleagues are using solenoid-based valves. “These valves are gentle enough to dispense cells without deformation, as well as being precise enough to have plate coefficients of variation around 5% or less.”
Technologies to try
As scientists automate more kinds of cell-based assays, more technologies will come in handy. As an example, Wehri says, “The acoustic liquid dispensers are very useful for getting accurate and reproducible amounts of small volumes of liquid.”
Editing the genome of a specific kind of cell can be used to study basic biology or in clinical applications. Here, Wehri explains, “CRISPR-Cas9 has made it a lot easier to make disease models in cells.”
To put cells in a more realistic environment, scientists often prefer three-dimensional cultures over the traditional two-dimensional approach. Organoids—3D cultures of cells that replicate at least some features of an organ—are increasingly popular, but not without hurdles to address. “The new challenges faced come about with the attempts to screen using 3D spheroids or organoids,” says Singhera. “Many organoid cell lines, once they form into a 3D structure, are much heavier than the surrounding media. As a result they tend to settle in approximately 15 to 30 seconds.” As Singhera adds, “The challenge comes about trying to dispense an equal number of spheroids per well especially in a 1,536 plate.” He and his colleagues are working on various approaches to these problems.
The software used to analyze results can really push ahead what can be done with many datasets. As Wehri points out, scientists can use artificial intelligence to “tease out more details from images both from ongoing campaigns as well as mining data you already have for lost treasure.”
Medical advances
In many cases, scientists apply cell-based assays to drug discovery and development. At Tecan, as an example, Magnan says, “We extensively work on setting up screening platforms in drug discovery for various therapeutic indications—such as oncology, metabolic disorders, or neurodegenerative diseases—using stem cells as well as 3D cellular models.”
In automating such applications, Tecan scientists use their newest generation of liquid handlers, FLUENT, which is designed as a highly versatile and user-friendly platform that facilitates integration of third-party devices. Based on a scientist’s needs, Tecan can design a solution for a specific assay and throughput. Tecan can routinely integrate a wide range of external devices—including incubators, shakers, stirrers, media heater and feeders, readers, high-content imagers and microscopes—and also provide a specific service, Labwerx, for customized solutions to address complex and “never done before” workflows, Magnan explains.
At The University of Virginia in Charlottesville, pathology professor Robin Felder and his colleagues design robots and develop automated cell-based assays, especially ones related to human kidney cells. Felder points out that “50% of Americans have hypertension or salt sensitivity.”
So, Felder collects live kidney cells from human urine, sorts them and runs cell-based assays. “From that, we can provide a personal salt index that tells you how much salt you can consume,” he explains.
For cell-based assays overall, Felder points out that a challenge is “maintaining a healthy environment for cells until they give you the information that you need.” For that, he says, “automation is essential.” He adds, “The fully automated 3D organoid-like cell culture process we developed controls the entire environment and cell manipulation so that cells are as close to in vivo conditions as possible.”
Image: Liquid handlers, like the Tecan FLUENT™, make it easier to automate cell-based assays. Image courtesy of Tecan.
There’s plenty of work left to do in automating cell-based assays. “Most of the automation is sitting in labs gathering dust, because the software is so hard to use,” Felder says. “If people could make systems turnkey then cell-based assays will really take off!”