Caitlin Smith has a B.A. in biology from Reed College, a Ph.D. in neuroscience from Yale University, and completed postdoctoral work at the Vollum Institute.
High-content screening (HCS) systems—which apply automation to high-resolution microscopy to look for changes in cells’ morphological or biochemical features—have become the workhorses of basic science and drug discovery labs alike. Coupling HCS with robotics, pharmaceutical companies can quickly screen large compound libraries, an integral step in the drug discovery process. Oncology, neuroscience and stem cell researchers, among others, use HCS to rapidly quantify the impact of pharmaceutical compounds or biological therapeutics on cells.
Today’s HCS systems are faster, more flexible, and more sensitive than ever. But not all systems are alike, and each has its pros and cons. Here we sample some current offerings, each with its own unique flavor.
Flexible and easy to use
Whether to save costs or bench space, many labs want major purchases to be multitaskers that can drive multiple applications. Molecular Devices’ ImageXpress® Micro XLS System fits this bill by serving both screening and research microscopy needs.
According to Jenny Hsu, senior marketing communications specialist in bioimaging at Molecular Devices, the system’s applications include “live-cell toxicity screening; wound healing; neurite outgrowth; 3D cancer-spheroid imaging; small-organism screening, including zebrafish, C. elegans and Drosophila embryos; [and a] homogeneous binding assay for antibody screening.” As an automated, high-throughput imaging system (up to 200,000 wells per day), its range also includes characterizing targets in key pathways, screening large libraries and phenotypic analyses of fixed or live cells.
A key component of the ImageXpress Micro XLS is the MetaXpress® analytical software. Turnkey application modules make more common analytical tasks easy to accomplish, though researchers can create custom modules and macros if they wish. But programming experience is not needed to use the software, says Hsu. “Whether users have little or advanced knowledge of image processing, the software provides an analysis continuum suitable for all experience levels.”
Analysis on the fly
Nikon Instruments’ High Content Microscope system, which couples the company’s Eclipse Ti microscope and NIS-Elements HC (High Content) software, allows users to analyze images “on the fly.” That means users can change acquisition plans or parameters during the experiment, if necessary, a considerable time-saver compared to waiting until after the experiment only to discover that you should have acquired the data differently. “This capability allows for immediate inspection during a plate run to get a sense of the data that it’s generating,” says Ned Jastromb, senior application product manager at Nikon.
Jastromb describes the Nikon HCS system as “the ultimate core-facility solution,” noting that it supports widefield, confocal, live-cell and fixed-cell imaging modalities. The platform is also applicable to both small-scale discovery experiments and large-scale screening assays, and users can transition directly from the former to the latter without adjusting such parameters as focusing method, wavelength, filters and camera settings.
Another characteristic that makes the system especially suitable for core-facility use is that “multiple focusing devices and options make the Nikon HC system agnostic to plate type and experimental design,” says Jastromb.
Nikon’s NIS-Elements software offers numerous built-in tools, including a “Plate View” tool that organizes images and analyzed data. But the software also enables researchers to create their own analysis strategies to account for their specific experimental parameters, says Jastromb. “Nikon provides the database infrastructure that can support the full recording of all device[s] and metadata, which is stored with each individual image.” On a more practical note, the software also can send researchers the results or experiment status by text or email.
Systems for different needs
Some labs more or less know what they want from an HCS system and have no desire to purchase features they will never use. GE Healthcare Life Sciences offers a range of systems to accommodate researchers with varying HCS needs. For example, the company’s simple-to-use Cytell Cell Imaging System is app-driven, rather like a tablet device. “[It] is ideal for users wanting to do simple routine analyses like a cell count, viability or cell-cycle assay, providing a simple entry-level system for personal use in the lab,” says Nick Thomas, principal scientist at GE Healthcare Life Sciences.
In contrast, the more complex IN Cell Analyzer systems provide greater analytical power and flexibility for advanced users with experience in designing their own analysis tools. But these also vary in complexity to cater to users’ different needs. For example, both the IN Cell Analyzer 2200 and the Cytell Cell Imaging System are widefield imagers with four-color fluorescence capabilities. The more advanced IN Cell Analyzer 6000 uses a laser-line scanning, tunable confocal system. This system “can be configured to optimize confocality in different color channels according to the user and assay needs,” says Thomas.
Separating spectral signals for greater sensitivity
Revvity introduced another confocal HCS system, the Opera Phenix™, in January 2014.
The company designed the Phenix to maximize speed without sacrificing sensitivity. Phenix relies on Revvity’s patented Synchony™ Optics technology, which controls excitation to reduce the “crosstalk” of fluorescent signals. “The confocal design, together with the automated water-immersion objectives, allows for effective optical sectioning of 3D micro-tissues, while the large field-of-view and the ability to simultaneously image up to four channels is helpful for phenotypic assays or rare-event detection,” says Jacob Tesdorpf, portfolio director of high-content imaging instruments and applications at Revvity.
The sensitivity of HCS systems can be limited at higher data rates. In particular, acquiring data while simultaneously analyzing data—which can save time in longer screens—sometimes limits sensitivity when fluorophore emissions interfere with each other. “The spectral overlap causes signals from one fluorophore with a shorter emission wavelength to be detected by the channel for the longer wavelength emitter,” Tesdorpf explains. For example, the broad blue emission bands of some fluorophores (such as the DNA stains DAPI and Hoecht) can overlap with green-fluorescent protein bands, limiting the sensitivity of assays that include nuclear translocation. Phenix’s Synchrony Optics separate the excitation and emission paths of fluorophores whose spectra overlap, minimizing crosstalk during simultaneous confocal acquisition by 98% on average, according to Tesdorpf.
Advances in HCS raise hopes that the technology will encourage breakthroughs in pharmaceutical screening using more physiologically relevant disease models. The heavy cost of drug candidates that fail because of unforeseen side effects are but one of many reasons that researchers are looking for new cell configurations or screening methods—for example, 3D cultures, microtissues and stem cell cultures. “Primary cells or induced pluripotent stem cell models require the ability to robustly describe and discriminate phenotypes,” says Tesdorpf. But can HCS by applied to complex disease models in 3D tissues? Advances in HCS technology may soon be equal to such complex tasks.