Featured Article
Wednesday January 20, 2010
by Catherine Shaffer
As more and more new assays are successfully adapted for high-throughput screening, the overall range of microplate-based assays has become broader. In addition to traditional fields, such as molecular biology, proteomics, and drug discovery and development, microplate instrumentation is showing up in areas such as food chemistry and environmental toxicology. Within the life sciences, microplate technology is used heavily for high-throughput assays, such as GPCR assays, kinase assays, ion-channel assays, cAMP, HTRF, SNP screening, and reporter gene screening. Microplates are also important in studies of protein-protein interactions or in detecting nucleic acids. Many cell biology studies are now done in microplates, in increasingly high volumes—cell growth assays, cell mobility assays, cytotoxicity assays, and, of course, cell-based drug screening studies.
It is an increasing challenge for microplate instruments to accommodate each unique protocol. A single protocol might include the use of an incubator, a vortexer, a liquid-handling robot, a PCR system, a microplate reader, or any of a variety of other types of instruments—each of which has to successfully handle the microplate. Newer technologies in microplate instrumentation offer ways of bridging the gaps between instruments and making the overall assay smoother, with fewer required human interventions. The widespread adoption of the Society for Biomolecular Sciences (SBS) microplate standards has made this process easier, allowing instruments to accommodate just about any microplate, or any microplate format. Newer technologies are also tailored to smaller wells, reducing errors that result from the extremely small sample size.
One instrument that is important, but often overlooked in equipping a laboratory for microplate-based assays, is the vortexer. Standard vortexers for microplates have a throw radius of 1.5 to 2 mm. This is barely adequate for a 96-well plate, and will not properly mix a 384-well plate. In order to create a vortex inside the well, the motion must overcome the capillary limit of the liquid. The Flurry vortexer from Stovall Life Sciences has a 1 mm throw radius, which allows effective mixing even of viscous liquids in 96- or 384-well plates. Since a SBS standard microplate well is 7.0 mm for a 96-well plate and 3.50 mm for a 384-well plate, the difference of half a millimeter is very significant. Says Ted Reynolds, president of Stovall, “Anything mixes 96, but 384 is difficult.”
When it comes to microplate readers, the market offers a wide variety. The types of assays supported include UV and visible spectrophotometry, luminescence, fluorescence, fluorescence polarization, and light scattering measurements. Some vendors also offer instruments that are tailored to specific assays, or specialized for high-throughput screening. Multifunction scanners have more than one detection mode, and multimode scanners offer additional tasks such as shaking and incubation. Some multiplate readers also incorporate liquid handling robotics, or are compatible with liquid handling modules that can be purchased separately.
Integrating all of these functions and formats into a single work flow is one of the most pressing issues in microplate instrumentation. This is the goal of the SiLA consortium, a group of system manufacturers, software suppliers, system integrators, and pharma and biotech corporations that are working together to develop standards for interoperability of laboratory automation system components.
The microplate reader company BMG Labtech has recently joined the SiLA consortium, and its product portfolio conforms to these shared standards in order to expedite the workflow for the end user. Their offerings include the PHERAstar series of multidetection plate readers for high-throughput screening, the Omega Series of modular upgradeable microplate readers with Tandem Technology, the NOVOstar advanced liquid handling microplate reader for cell-based assays, and the NEPHELOstar for compound solubility screening. “The challenges, of course, are different LIMS platforms, and different data formats. Many companies have their own requirements in terms of how the data should look before it goes into the LIMS system for further software evaluation.... Data and hardware drivers should be standardized and everybody should then adhere to this format, so that different devices like microplate readers, incubators, and other robotic platforms speak the same language,” says Dr. Michael Fejtl, international sales and marketing specialist with BMG Labtech.
For customers and as a general resource center, BMG Labtech maintains an extensive library of application references. In one of more than 1700 references in the library, scientists from Chapel Hill North Carolina make use of the POLARstar Omega for FRET-based screening for potential modulators of the Galpha-i1 protein/GoLoco interaction. The results demonstrate a robust assay using CFP-YFP FRET, with reliable performance from the POLARstar Omega and the included MARS Data Analysis Software. Because microplate assays can produce a large volume of data, powerful data analysis software like MARS is vital, and portability of data is an important goal of BMG Labtech.
The increasing variety of microplate assays can be a challenge for individual researchers, as well as for vendors serving the larger market. Budget limitations can force painful choices in instrumentation. Newer, modular multimode plate readers can take some of the pressure off of a researcher that has a need for more than one microplate device.
Multimode microplate readers are used for a variety of absorbance, luminescence, and fluorescence-based assays. Fluorescence detection in particular is used in hundreds of different assays, and over a wide range of wavelengths, for basic research as well as drug screening applications. As a result, monochromator-based systems are ideally suited for these applications with their flexible wavelength selection system. However, monochromator-based optical systems have limitations that hinder the performance of some assays. Examples include fluorescence polarization, AlphaScreen/AlphaLISA assays, as well as any fluorescent assay that would benefit from a larger excitation and/or measurement bandpass.
Filter-based readers don’t have this limitation but are a lot less flexible, since each fluorophore requires a dedicated filter set. They also don’t allow spectral scanning. Until recently, users had to make a decision upfront (filter vs. monochromator) when purchasing a multimode microplate reader, which limited the application range of their new instrument. The new Synergy™ H4 from BioTek Instruments combines both detection systems (monochromator-based and filter-based) in one patent-pending Hybrid system to address the large diversity of fluorescence-based assays, so that it is not necessary to choose between a filter- or monochromator-based system. Additionally, modular read modes can be added individually. Says Xavier Amouretti, product manager, “The Synergy™ H4 and the entire range of Synergy™ plate readers are modular, so a customer only purchases exactly what they need. As additional assays are added to their research, the reader can be easily upgraded so that there is no interruption to their workflow.”
Researchers are now beginning to reap benefits from early, difficult efforts to standardize microplates and microplate instrumentation. It is now quite simple to swap microplate components in and out of an automated laboratory workflow, as well as porting data from one system to another. This reduces labor and wasted time trying to translate systems and data from one “language” to another.