© 1998 by Oxford University Press
The race to isolate single cells for genetic and molecular analysis has grown contentious, with a rift as broad as the Atlantic and as thin as a laser beam separating the three competitors. A technology developed at the National Institutes of Health, called laser capture microdissection (see News, Dec. 4, 1996), dominates the small, but growing, United States market. However, two other companies -- one in Germany and the other also in the United States -- are actively criticizing the NIH system and proffering their own as superior.
So far, the stakes are relatively small. The companies each report worldwide sales of only 40 to 100 instruments, which wed lasers to microscopes and zap small groups of cells into test tubes. But within a decade, the expectation is that these instruments will work their way out of the lab and into the clinic as staples for cancer diagnosis and care. Whoever perfects the best system will be poised to reap huge financial rewards.
"Just like everything else we do in technology, there are going to be different solutions and let the best solution win," said G. Steven Bova, M.D., a pathologist at Johns Hopkins University who uses the NIH system. "It will all boil down to the usual things, which are cost, convenience, and proven effectiveness."
Two years into the contest, the three systems cost about the same, upwards of $80,000. But convenience and effectiveness are disputed. A thin stack of journal articles, press clippings and sales brochures make up the whole of printed material on laser microdissection -- hardly enough to make an informed choice. Several researchers said they are curious about the technologies but want to know how well each of the systems work.
"For the near term, the whole process of microdissection . . . needs to be scientifically tested and peer-reviewed," said Tom Baer, Ph.D., cofounder and chief executive officer of California-based Arcturus Engineering, Inc., which licenses and manufactures the NIH-developed system.
The NIH-Arcturus system, called PixCell, appears to have the marketing edge. It has been published in more prestigious journals, conjured more popular press coverage, and generated better name recognition, at least in the United States. The system from the German company, PALM (Positioning and Ablation with Laser Microbeams, an acronym that also works in German), reigns in Europe, an area Arcturus is also eyeing. New Mexico's Cell Robotics, Inc., recently entered the fray by marketing to the same niche Arcturus and PALM are after -- cancer genetics labs.
Several areas of dispute, including precision, purity of samples, and type of laser used, provide raw material for a fracas unfolding at conferences and in journals. One of PALM's lead researchers, Karin Schütze, Ph.D., co-authored an article in the September Journal of Molecular and Cellular Biology touting the superiority of PALM's technology. Another offering from Schütze, in the August Nature Biotechnology, drew coverage from Science News and similar outlets, which hooked into the claim that PALM had isolated single cells for DNA amplification and analysis. Though couched in parabolic science-speak, this article also found flaws in PixCell's approach.
Baer sees these attacks as unfair. "What we're being criticized for is untrue, and the things that are even slightly true we have solutions for," he said. He added that PALM's aggressive stance does not phase him, but that it bothers members of the NIH team.
When asked about the effectiveness of the PALM system, Baer said he doesn't think it is "appropriate to comment publicly on a competitor's technology. Customers should really judge the benefits of each technology on its own."
Difficult to Judge
And what are the benefits of each? It depends, of course, on who is talking. The University of Wisconsin's John White, Ph.D., is perhaps as unbiased a voice as one finds in the relatively small field of laser microscopy. White is not linked to NIH or any of the companies involved, and he uses lasers for neurological, not cancer research. But even he has heard noise from the scuffle.
"Arcturus's marketing has taken a fairly aggressive commercial twist, so it's hard to know how well [their system] works," he said. When asked to explain, White said a company he consults for was goaded into supporting the PixCell system, and that he had trouble duplicating the technique. The critical problem: he could not get target cells to stick to a piece of film as required. Granted, White was not using the same setup Arcturus sells. "They [Arcturus] may have some magic formulation of the glue [film] that works every time," he said.
Julie Foley, who works in the National Institute of Environmental Health's Experimental Pathology Lab, said sometimes PixCell requires more work than advertised. For instance, in one type of study, called loss of heterozygosity analysis, users "need to fire the laser like 5,000 times. You'd get carpal tunnel syndrome trying to do that."
Other field scientists give more positive reports. Steve Reynolds, Ph.D., a molecular biologist at the National Institute for Occupational Safety and Health in West Virginia, was delighted with his new Arcturus machine, which he tested under ideal conditions with guidance from Arcturus engineers. "It's so well designed, we got it to work on the first try," he said. "The initial feeling was I missed something. It seemed too simple."
Ignacio Wistuba, M.D., at the University of Texas Southwestern Medical Center, Dallas, is also happy with his Arcturus system. But he relies on an older manual microdissection method about half the time, using needles, dexterity, and patience to poke cells into test tubes. "The problem with the machine is that the laser is still too big. It's good, fast, clean to use on big tumors, but it's not very precise for the smaller ones."
A Tale of Two Lasers
Wistuba's complaint focuses on perhaps the most contentious point of the microdissection debate. Arcturus' infrared laser spreads to a width of 30 microns, covering about three cells. In contrast, PALM's ultraviolet laser tightens to a sub-micron focus. PixCell works like a hole punch, capturing a circle of cells with each laser burst, while the PALM system slices around the targeted area like a welder's arc.
These differences could impact the purity of procured samples, which is critical for gene identification, the goal of microdissection projects like the Cancer Genome Anatomy Project (see News, Dec. 18, 1996). However, it is clear that both laser technologies outperform older manual methods like those used by Wistuba.
Before lasers, scientists scraped ensembles of cells -- some cancerous, some not -- and jammed them into test tubes, jumbling the cells' DNA. This jumble made identification of potential cancer genes difficult -- like trying to listen to a single bird in a cacophonous forest.
"If you have thousands of genes on a chip [also called 'microarrays'; see News, Dec. 18, 1996] and you want to analyze tissue with that, if you just grind it up and put it on the chip it's meaningless, no matter how sophisticated the chip is," said Lance Liotta, Ph.D., a National Cancer Institute scientist and part of the NIH team.
That's why researchers "want to be able to capture single cells, to analyze as little material as possible," added Foley.
Baer calls single cell analysis "a kind of holy grail," and it appears both PALM and the NIH-Arcturus team have their hands on the cup. In their Nature Biotechnology article, Schütze and co-authors describe isolating single cells from a slice of a colon adenocarcinoma. And Liotta says scientists routinely isolate large single cells with Pix-Cell. But such a spectacular feat may not be necessary or useful.
"It's great if you can get a single cell," said Nicole Simone, who helps train researchers on PixCell at NIH's microdissection lab. "But what can that really tell you? Maybe you'll get an anomalous cell that won't reveal much about the [tumor] area you're looking at." A paper by Simone and Liotta supporting this assertion is under review at a scientific journal.
Another problem: Analyzing the tiny bits of DNA from single cells is tricky, and has some, like Foley, skeptical about the usefulness of single cell capture. "The hard part is learning how to tweak the molecular analysis and amplification, because you're working with fewer cells than usual," she said.
"From a pathology point of view, from an actual practice of medicine point of view, it's not clear that single cell extraction is going to be critical for a diagnosis," said Baer. "But to propel the research forward, I think that understanding the subtle differences between cells is a critical advance."
A Serendipitous Market
PALM's system, like Cell Robotics', springs from 30 years of research aimed at performing surgery inside cells; much of this research focused on in vitro fertilization. That their systems can detach cells from one another is a side benefit, not an originally intended function. Such versatility comes from using an ultraviolet laser, which runs at shorter wavelengths, and hence higher energy, than PixCell's infrared beam. But ultraviolet's precision comes with a price: the danger of damaging DNA or other parts of cells.
"I'd sure as hell prefer to have an infrared laser," said Wisconsin's White. "Infrared tends to be much less damaging to the cell."
Although it's well-established that ultraviolet light breaks molecular bonds and harms cells, at any given firing there's no way to tell if any damage will be done. "There's a risk you take with ultraviolet lasers," said White. "You don't know if any of the photons will hit any critical [molecular] bonds, so it's just a matter of how much risk you're willing to accept."
With this potential problem in the fertilization market (prospective parents may balk at even a slight possibility of DNA damage), and with a product to make profitable without government help, PALM and Cell Robotics turned their ultraviolet lasers toward microdissection.
In contrast, PixCell was specifically developed for tissue microdissection and eventual clinical use. When developing their system, Liotta and NIH partners Robert Bonner, Ph.D., National Institute for Child Health and Development, and Michael Emmert-Buck, M.D., Ph.D., National Cancer Institute, maintained ease of use as a priority.
It seems they succeeded: Training researchers on PixCell, even those new to microdissection, takes but a few hours. The PALM instrument is trickier; a company representative told an interested conference-goer that it would take a few weeks to get the hang of their system. PixCell has also been field tested more thoroughly, as shown by its use to identify a gene implicated in pancreatic cancer, called MEN1.
Beyond speeding identification of genes implicated in cancer, laser microdissection's advantages should hasten its adoption as a diagnostic tool. Such a tool -- no matter which company it comes from -- will increase patients' survival chances by generating genetic profiles of tumors, a few cells at a time. Liotta said doctors will be able to use this information "to predict the outcome and the best treatment for cancer patients."
-- Brian Vastag
How to Slice It? Tissue Technologies Vie for Potentially Lucrative Market
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