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Jul 1, 2009 (Vol. 29, No. 13)

Upgrading Cell-Based Viability Assays

Novel Approaches More Accurately Reflect the In Vivo Human State

  • Molecular Probes

    Invitrogen, a division of Life Technologies, has available through its Molecular Probes brand a wide variety of options for assaying cellular viability and cytotoxicity. These encompass a number of fluorometric assays that possess significant advantages over traditional colorimetric and radioactivity-based approaches.

    Mike Janes, manager of R&D for Invitrogen’s cell health assays business, and George Hanson, Ph.D., principal scientist for discovery assays and services, elaborate on the company’s products, comparing automated high-content cellular imaging with high-throughput screening technologies.

    The two strategies are appropriate for different types of investigations, since cellular imaging allows an overall look at a mixed population of cells, but lacks the speed of high-throughput screening. While some Invitrogen products employ classical approaches to measure general cell viability and cytotoxicity, the Click-iT® platform is the basis upon which new assays for apoptosis and nascent synthesis of DNA, RNA, or protein have been developed. 

    According to Janes, Invitrogen’s development of capabilities to investigate perturbations of lipid metabolism is a recent new direction for the company. The HCS LipidTOX™ kits are designed for the detection of phospholipidosis and steatosis; they offer a complete solution for performing image-based high-content screening assays. The toxic side effects of disrupted lipid metabolism can be triggered by various foreign compounds. The probes are available in a range of colors, detectable with fluorescent microscopes and quantifiable with image analysis software available on HCS readers.

  • Multiplexed Assays

    Promega offers a number of cell-based assays based on novel bioluminescent and fluorescent chemistries that can be combined in the same well, providing more biologically relevant information per well of cells. Moreover, the newer assay chemistries are easy to implement on robotics platforms and scalable up to 1,536-well plates, according to Pam Guthmiller, strategic marketing manager for cellular analysis.

    “By developing assay chemistries that can be multiplexed in the same well, the researcher can now perform these assays in microwell plates using a simple multimode detection instrument. Because of the simplicity and sensitivity, a number of life science researchers are turning to these formats.” 

    The offerings are versatile and sensitive, according to Guthmiller, and include the GloMax®-Multi detection plate reader system for measuring a broad range of fluorescent and luminescent cell signals. Many of the assays are streamlined through a homogeneous add-mix-measure format, meaning that removing or changing the medium is unnecessary. The various substrates are simply added to the culture medium as a concentrate and then read. Examples include the MultiTox-Fluor multiplex cytotoxicity assay (measures viability and cytotoxicity) combined with the Caspase-Glo® 3/7 assay, designed to provide data on the role of apoptosis in cell cytotoxicity.

    These assays can also be combined with reporter assays to gain information on gene expression when cells are exposed to potential therapeutic drugs. The ONE-Glo™ luciferase reporter assay system can measure transcription and translation. Combined with the CellTiter-Fluor™ cell-viability assay, evidence can be assembled on the concordance between gene expression analysis, cell viability, and cytotoxicity within a single assay.

    Another key product is the nonlytic, luminescent CytoTox-Glo™ cytotoxicity assay. “It can detect small changes in viable cell number and also can be multiplexed with fluorescent-based assays, improving data quality and saving time, money, and reagents,” says Guthmiller.

Readers' Comments

Posted 07/10/2009 by Sr. Director, Business Development

Very insightful article that highlights multiple new technologies being advanced. One concern that I do not see being addressed is the drive to make our in vitro assays more physiologically relevant by removing and replacing non-human components when possible. Cells do b ehave differently when cultured on plastic or Matrigel when compared to fully human ECMs. There are dramatic morphological as well as molecular gene expression differences that lead to aberrant and misleading results. Minimizing these impacts on in vitro assays and maximizing human physiological relevance is the future of cell-based assays.

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