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.
Performing molecular and cellular analyses in multiwell microplate formats is an important methodology for high-throughput screening, or for boosting throughput generally. In addition to increased throughput, microplate formats enable researchers to miniaturize their analysis (using less sample and reagent) to study regulatory events and biochemical reactions. Today’s multimodal readers offer an array of features for detection, measurement and analysis of samples in microplates. Here we review some of the latest options in microplate readers available for researchers.
Measurements
For years, multimodal microplate readers reliably delivered measurements of fluorescence intensity and absorbance. Today many readers include luminescence measurement as a standard feature. Some researchers are perfectly satisfied with these options, but additional detection modalities include: flash luminescence (using injection prior to measurement), time-resolved fluorescence, fluorescence resonance energy transfer (FRET), time-resolved FRET, fluorescence polarization (TR-FRET), fluorescence spectral scanning, Alpha technologies (such as AlphaScreen®) and bioluminescence resonance energy transfer (BRET).
Modularity
Some microplate readers are designed to be modular so researchers can add functionality to the instrument as needed. Examples of new upgradeable systems include Berthold Technologies’ TriStar2S LB 942 microplate reader, Molecular Devices’ SpectraMax® i3x Multi-Mode Microplate Reader, BioTek’s Synergy™ Neo 2 and Cytation™, which can also be upgraded to add imaging and Thermo Fisher Scientific’s Varioskan™ LUX multimode microplate reader. “Due to its modular concept, the functional range and the measurement technologies of the TriStar2S LB 942 are expanded regularly,” says Frank Schleifenbaum, head of product management and business development, bioanalytical instruments, at Berthold Technologies. The TriStar2S LB 942 uses a patent-pending Berthold PHA-detector for background-free fluorescence measurements that are tolerant of temperature fluctuations.
Western blot detection
Molecular Devices offers the ScanLater™ Western Blot Detection Cartridge, which enables researchers to detect and quantify Western blot signals using the SpectraMaxi3x Multi-Mode Microplate Reader.
Dynamic range
Dynamic range represents the measurement of the smallest to largest statistically significant detected signal that an instrument can collect.
Having a wider dynamic range can potentially provide more sensitive and accurate detection as well as preservation of sample.
The Synergy™ Neo2 Multi-Mode Reader from BioTek instruments combines ultra-fast read speeds while still maintaining high sensitivity; with an extended dynamic range of seven multdecades for fluorescence intensity measurements, and greater than six decades for luminescence measurements, along with a host of other flexible reading modes.
In the case of Molecular Devices’ SpectraMax i3x Multi-Mode Microplate Reader, the device has a reported dynamic range of at least six decades. This can “eliminate the need to dilute or concentrate precious samples,” notes Cathy Olsen, application scientist at Molecular Devices.
Cell imaging
Several microplate readers now include cell-imaging capabilities. Tecan’s Spark series of microplate readers offers researchers the chance to use both monochromators (to define an exact excitation wavelength, for example) and filters (to define wavelengths while enabling greater sensitivity for detection, for example) in one experiment. “This helps to reduce time for assay development and saves costs for precious samples and cell lines,” says Siegfried Sasshofer, director of marketing detection at Tecan. BioTek Instruments’ Cytation™ Cell Imaging Multi-Mode Readers integrates digital imaging, including brightfield, color brightfield, phase contract and fluorescence microscopy, into a compact unit along with filter- and monochromator-based detection for data-rich qualitative and quantitative, and offer temperature and environmental control for real-time imaging during cell-based studies or other sensitive workflows. Other readers with cell imaging include Revvity’s EnSight™ Multimode Plate Reader and Molecular Devices’ SpectraMax i3x Multi-Mode Microplate Reader.
Label-free detection
The advantage of using labeled probes is detection specificity. However, when a specific probe is not available, or it causes off-target effects on the cells being analyzed, researchers often turn to label-free detection to overcome these challenges. Revvity’s new EnSight Multimode Plate Reader reads label-free Corning® Epic® technology (in addition to standard labeling technologies) and is appropriate when multiple assay types are more important than speed. “The EnSight reader is chosen for orthogonal assays and for multiparametric results, since it can combine results from microplate imaging, label-free technology and labelled technologies within one assay approach,” says Volker Eckelt, portfolio director, multimode detection, at Revvity.
Molecular Devices’ SpectraMax MiniMax Imaging Cytometer is an optional feature that lets researchers count cells and monitor cell growth using the company’s StainFree™ Cell Detection Technology (in addition to fluorescence imaging) on the SpectraMax i3x Multi-Mode Microplate Reader. Tecan’s Spark readers also feature label-free cell-counting capabilities.
Incubation
Tecan’s Spark readers are optimized for more biologically relevant assays, says Sasshofer. “We’ve had an especially big demand for microplate readers that support and enhance cell-based assays,” he says. The Spark readers’ Te-Cool™ cooling module can maintain the interior temperature below ambient levels. “We believe this is important because chemical reactions are highly dependent on temperature, so even small temperature changes from day to day or lab to lab can lead to variability,” says Sasshofer. “Holding the temperature steady can increase the consistency of the results.”
The Synergy™ Neo2 and Cytation™ Multi-Mode Readers from BioTek Instruments offer integrated controls, including gas, temperature and shaking, to optimize the chamber environment for cell-based assays, and can be linked to the company’s BioSpa 8 Automated Incubator to facilitate walkaway automation of long-term live cell assay workflows.
Throughput and automation
Automation is one of the most effective ways to boost throughput when working with microplates. The addition of plate stackers supplements the sample throughput while also providing walk-away convenience for researchers. Automation-compatible readers are available from many vendors, including Berthold Technologies, BioTek Instruments, Molecular Devices and Thermo Fisher Scientific.
Multiplexing measurements is another way to increase throughput. Revvity’s EnVision® Multilabel Reader, which reads up to 3,456-well microplates, can multiplex dual emissions in AlphaPlex or BRET assays, for example. “Both readers feature the unique HTS-Alpha and ultra-sensitive luminescence technology, with an ultra-sensitive photomultiplier tube due to the contact aperture,” says Eckelt. “It’s ideal for even ultra-high-throughput screening.” Molecular Devices’ SpectraMax i3x Multi-Mode Microplate Reader and Thermo Fisher Scientific’s Varioskan LUX reader also perform multiplexing.
Ease of use
Because microplate readers are meant to shoulder routine and time-consuming tasks for researchers, ease of use is an important feature.
Automatic settings, for example, “can simplify the workflow and ensure accurate results without the need to manually adjust settings,” says Olsen. For example, the new Thermo Scientific™ Varioskan LUX reader’s automatic dynamic-range feature “selects the optimal reading range based on signal intensity in the well and ensures high sensitivity and wide dynamic range, even if you have samples with both very high and low signals in the same plate,” says Reija-Riitta Harinen, product manager at Thermo Fisher Scientific.
Promega’s newest GloMax® series of microplate readers — including the Discover, Explorer, and Navigator systems — automatically adjusts the gain of the instrument when measuring signals, and has little well-to-well cross-talk. “When you have neighboring samples in the plate, you can be confident the signal you detect is from the particular well of interest and not from the neighboring well,” says Michael Bjerke, Promega’s global product manager.
Adyary Fallarero, lecturer and adjunct professor in the Division of Pharmaceutical Biosciences, Faculty of Pharmacy, at the University of Helsinki, likes the automatic dynamic range feature, saying that it streamlines “the development of new assays and optimization of assay conditions.” Fallarero currently uses the Varioskan LUX reader for measuring fluorescence, absorbance and bioluminescence signals in mammalian cell-based assays. “In many cases, we perform quantification with the Varioskan LUX, and fluorescence imaging using the Evos Fl microscope, in the same microtiter well plate,” she says. “We have also performed enzyme-based kinetic assays with automated injection, and spectral scanning assays.”
Automatic features save time and can even help prevent accidents. For example, the Varioskan LUX reader incorporates safety controls that alert researchers to oversights such as a missing microplate or misplaced reagent-dispenser head—heading off spills before they happen.
Software also is becoming simpler and easier to use. For example, Tecan uses an app-based approach for routine assays such as DNA quantification, quality checks and cell counting. “Researchers want access to cool features without software hassles,” says Sasshofer.
Although it’s easy to be dazzled by the sophisticated features on today’s microplate readers, the most critical feature to consider is whether the instrument is a reliable instrument that will increase your work throughput while providing the most accurate data. If you focus on what you are likely to need, it should be simple to find a multimodal microplate reader that supports your research both now and into the future.
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