By Laura Lane
Let’s face it: Researchers need to be treated like the sophisticates that they are. No more table wine. Generalized biochemical data simply doesn’t cut it anymore. Neither does molecular information without a hint of time and space. More and more, life scientists need the nit-picky, vintage-level details of cellular imaging data.
For years, labs subsisted on assays that revealed interesting information on the kinetics and biochemistry taking place in cells. Technology now allows researchers to merge those findings with the spatial and temporal data that solidifies concepts of cell physiology and gene expression. Imaging now means viewing the location of specific molecules within a cell and the surrounding structures. Melded with time-lapse approaches, microscopy reveals cause-and-effect relationships and allows researchers to follow the physiological processes from genetic loci to the cytoplasm.
“No longer are components of chromatin, nucleosomes and the transcriptional machinery merely protein bands on a Western blot or fluorescents dots in a fixed cells, but can now be followed as they actively assemble and interchange at gene loci and sites of transcription,” wrote the authors of a paper published in a 2006 issue of The European Molecular Biology Organization Journal.
The technology to do so has arrived just in time, as researchers realize the limits of data culled from cell populations. Analyses based on averages can mislead. The issue is especially stark with tissue specimens, such as tumors and the heterogeneity of its cells. Some reveal the smoking gun, but nearby cells can come up negative. Taking the stochastic approach of comparing individual cells can thus help researchers understand the genetic and other differences of normal and diseased cells.
In the simplest of terms, the new technology is just an extension of the traditional, modified to support drug discovery efforts, says Baggi Somasundaram, product manager of Bio-Imaging, BD Biosciences, which sees its strength as offering complete solutions – including instruments, software, consumables, reagents and support – to address applications for high content cellular analysis. With instruments, the company’s approach is to offer integrated imaging systems that provide automated image capture and quantitative analysis of cell populations and their sub-cellular components.
“We’re basically reinventing the microscope for drug discovery,” Somasundaram says. “We’re taking the microscope and automating it.”
Researchers are eagerly diving in, acquiring the newest devices that reveal the detailed events of the cellular world. Biocompare’s latest market report, “Imaging and Analysis Tools: An End-User Survey,” published in September 2007, confirms researchers’ acceptance of new imaging technologies. The survey shows an upward climb in sales for the latest confocal, high content, and molecular imaging equipment and, according to the report, “reveals a healthy market enthusiastic about and educated on new features and advances in capabilities.”
“We’re seeing an explosion of technology and also an explosion of desire,” says Richard Levenson, director of research for biomedical systems at CRI, Inc.
The advances are big and pertinent enough that researchers are indeed ditching older equipment for the new generation of instruments. The instruments include those that feature automation, which was once developed only for big pharmaceutical companies, at a scale and price suitable for individual labs. Devices for multiplexed, high content, and high throughput protocols continue to draw interest, especially now that they come with software that is truly geared toward the average researcher.
“More and more people, who are not microscopists, are being put in front of a microscope,” Levenson says. “They want to quickly generate a tool to help analyze something without having to involve an expert.”
The first step could be making the right purchase. Unfortunately, the task is ever more confusing, with all the enthusiasm over fancy instruments that show more, tell more, and do more. But, if you’re only looking at cells in a dish or very thin slices of tissue, you may only need a standard light microscope. “Nine out of 10 times you don’t need a confocal microscope if you’re not asking very specific spatial questions,” Levenson explains.
To integrate multiplexing, you need only purchase CRI’s Nuance, a cooled charge-coupled device that fits on most any microscope. Inside, a tunable filter records various wavelengths, which enables the creation of an image from each wavelength. Called multispectral imaging, the Nuance allows researchers to simultaneously use various colored dyes without the confusion of overlap.
“Multispectral imaging makes sure you’re looking at what you’re looking at,” Levenson says.
While multispectral capabilities offer insight into layered colors, multimodal imaging allows the examination of cells within live animals in layered formats, including fluorescent optical visualization, radioisotopic detection, and x-ray. Whereas many whole animal imaging systems depend on a dedicated technician or a physician skilled in magnetic resonance or computed tomography, the Kodak In-Vivo FX Pro by Carestream Health, Inc. can be operated by any researcher, says Bill McLaughlin, director of research and development of molecular imaging at Carestream Health. The accompanying software may also come as second nature. Its design stems from well-developed programs used for analyzing gels and blots.
Having all three types of modalities wrapped up into one “is really important because you can take them all without moving the animal,” adds McLaughlin, who says that the company introduced a model with multispectral imaging this year. “The images are all taken in the same focal plane that overlay precisely on top of one another, so you get a much better idea of where molecular signals from targeted cells are located based on the X-ray based anatomical structure of the animal."
That’s the idea with TTP Labtech’s Acumen eX3, but with single cells. Functioning as a high content system, the instrument opens the door to the efficient screening of adherent cells growing on the bottom of 96-well plates. Flow cytometers served as the earliest high content screeners but could only be used for non-adherent cells, says Wayne Bowen, chief scientific officer at TTP LabTech. The new technology gives a boost to oncologists who work with transformed cells, most of which are adherent.
The Acumen eX3 requires the use of special multiwell plates designed with a clear bottom and black walls for each well. A laser scans the cells from the bottom, detecting fluorescent signals. A series of imaging sessions allows detection of proteins that have changed position within the cell, such as from the nucleus to the cytoplasm.
“There no change in total fluorescence,” Bowen says. “You just see the fluorescent signals moving from one place to another.”
