This is the second of a three-part series on the emerging technology. If you haven't already done so, you might like to begin with Part 1 of this series.

Also in this article:

• Cepheid bridges the sample-volume gap
• Microarray technology development at Genometrix
• Caliper harnesses electrokinetics
• Vysis prepares to launch Genosensor array technology
• Microtechnology development at PE Applied Biosystems

For those who marvel at how quickly DNA chips have burst upon the scene, prepare to be astonished further. Most companies report rapid progress, and suggest that as more is learned and applied, the pace of development in all fields of genetic-related diagnostics and therapeutics—including DNA microarrays—will continue to accelerate.

Labchip by Caliper Technologies (Palo Alto, CA), showing photolithographically etched fluid channels (in red). Channels are typically 70 µm wide and 10 µm deep. Photo Courtesy Caliper Technologies

 

There is no better example of this than the Human Genome Project (HGP). When the project was launched in 1990, government officials viewed it as perhaps a 20-year project. Proponents compared the vision of the project to that of America's goal of landing a man on the moon in the 1960s—but conducted with international cooperation and over an even greater period. Almost as soon as the HGP was launched, the date for completion was revised to 2005, then to 2003, thanks to developments in computerization, telecommunications, and molecular biology. Today, because of DNA chips and related developments, some officials now believe that the HGP can be completed by the end of this decade—three years ahead of even the revised schedules. In turn, as more is learned in this massive effort, it will accelerate DNA-chip development even more, observers say.

This is the second installment of a three-part series on DNA-chip technologies. The first reviewed the theoretical underpinnings of the field and examined the market forces driving product development. This installment will look at the state of the art and the various competing technologies in this embryonic field. The final article will cover the challenges to commercialization facing companies engaged in this new marketplace as well as prospective near- and long-term applications for these technologies.

Fast-Moving State of the Art

"The outlook is changing fast," says Deepak Thakkar, manager of microarray products at Genometrix (Woodlands, TX). "As an industry, we have largely been in a hibernation period, but in the past 18 months we have introduced a lot of highly targeted products based on very specific needs."

As an indicator of how rapidly the pace of development has proceeded, consider that the first DNA chip, by Affymetrix (Santa Clara, CA), was introduced only two years ago. Today two dozen or more firms are actively engaged in developing microarray technologies, and many others are developing related technologies for sample preparation and analysis.

"Until recently, most of this development work has been centered on what I would broadly call detection," says Peter Wilding, PhD, director of clinical chemistry and professor of pathology and laboratory medicine at the University of Pennsylvania (Philadelphia). "The vast majority of the products that have been introduced, however, are sold for highly specialized applications, usually in research-laboratory or drug-development applications."

Such applications usually involve some genomic-related problem. For example, the gene p450 and its eight different expressions, or mutations, have been linked to various cancers. Researchers are examining exactly which forms have the closest link to these cancers. In turn, drug developers want rapid genomic results so they can quickly determine the effectiveness of candidate therapies.

The reason that research-laboratory and drug-development applications are being developed is simple: that's where the money is. Governments, drug companies, and universities have poured billions of dollars into developing DNA chips for applications related to their agendas.

Take governments. The Defense Department's Advanced Research Projects Agency (DARPA) and the Commerce Department's Advanced Technology Program (ATP) at the National Institute of Standards and Technology (NIST) have invested millions of dollars as seed money for developing both various kinds of chips and their basic manufacturing technologies. "All the major players in this field have received research money from ATP," says Uwe Müller, director of advanced technology at Vysis, Inc. (Downers Grove, IL). "Stan Abramowitz and NIST should get a lot of credit. The ATP is a government program that is actually working, even with minute funding levels, and its support of this technology is the envy of the world."

DARPA has contributed millions more. In its case, DARPA funds projects for "dual use," meaning they must have both civilian and military applications. The prospective military use of DNA-chip technologies is for advanced detection of battlefield biological and chemical weapons.

Private-sector sources of funding have followed with billions more, initially from venture capitalists, and later from strategic partnerships, institutional investors, and public stock offerings. Yet these sources of funding have made pursuit of short-term financial objectives even more urgent, several observers point out.

As an example of this, Müller recalls: "Before Vysis was spun off from Amoco Technology, we were working on a food diagnostic application for E. coli testing; but we put aside that project. We realized that the food processing industry is very competitive, and the low profit margins for the tests used in that industry would not bear the heavy costs of research and development for DNA chips."

Today, such economic inducements are readily apparent. Almost every major manufacturer of DNA chips has a strategic relationship with a major drug company, taking advantage of the eagerness of pharmaceutical manufacturers to find ways to screen potential drugs faster and more accurately. These relationships usually consist of "early access partnerships," in the words of Lewis Gruber, president of Hyseq (Sunnyvale, CA). "This means that partners have access to a company's technology, often for specific applications only, before others do."

"If these technologies succeed in giving pharmaceutical companies the ability to screen drugs quickly and accurately, the payback will be tremendous," agrees Thakkar. "Only one drug in 10,000 succeeds in the marketplace and it takes 12 to 15 years to develop each new one. If these technologies can reduce the length of development time to eight years, that translates into significant savings for these companies."

An example of such corporate relationships is the set of strategic agreements announced earlier this year by Affymetrix and Eos Biotechnology, Inc. (South San Francisco, CA). The agreements allow Eos broad access to Affymetrix's custom and standard GeneChip expression chips for Eos's molecular genomics research efforts in specific fields of cancer, inflammation, and cardiovascular disease.

However, some analysts, including Peter Wilding, believe that such drug and research partnerships are stifling innovation that could lead to larger markets. "If the use of chips remains an activity that is confined to research laboratories—because you need their equipment to prepare samples and do the analysis—then the markets for DNA chips will be limited," he says.

Such thoughts are leading companies into the next phase of microarray development, which is already well under way. A number of companies are developing technologies that use immunoassay reagents or amplification techniques such as polymerase chain reaction (PCR) to prepare samples right on the DNA chip. Companies are also exploring technologies for conducting sample analysis on the same chip and outputting results to a single small instrument. Taking small sample amounts and moving them through the various stages of sample preparation, detection, and analysis of test results is being made possible by advances in capillary electrophoresis. Using electrical charges and micro- and nanoscale structures, researchers have successfully demonstrated that it is possible to perform all of these stages on a single DNA-chip system.

"We think we can have such a product in the marketplace by the third quarter of 1999," says Cris McReynolds, director of business development for Cepheid (Sunnyvale, CA), a maker of sample-preparation and analysis equipment designed to be integrated with DNA-chip technologies. Others echo his comments regarding their firms' product development timetables.

Types of DNA Chips

There are three basic types of DNA chips. The first and oldest is the sequencing chip. This is also the type most commonly discussed in popular articles about this technology. With sequencing chips, such as those initially produced by Affymetrix or Hyseq, segments of DNA (usually 20 bases long) are placed in a microarray. Target samples are then introduced to the chip and the segment that the sample "sticks to" (or hybridizes with) determines the result. This design is called sequencing by hybridization (SBH), and is both an industry term and an intellectual property of Hyseq, says Gruber. Many other companies are now producing sequencing chips, most using the SBH approach. But whatever their technique, such products are intended to determine the DNA sequence of the sample.

The second variety of DNA chips is known as the expression chip. These are designed to determine the degree of expression of a certain genetic sequence by measuring the rate or amount of messenger ribonucleic acid being produced by the target gene. This is done by creating chips with a specific set of base pairs (as opposed to sequencing chips, wherein every possible base-pair combination is arrayed). Results are then compared to a reference or control, and the degree of change is noted. These chips are useful in diagnosing and treating diseases linked to particular genetic expressions, such as some forms of cancer. Vysis and Synteni (Fremont, CA) are two companies engaged in marketing expression-chip-based products and services.

The third type of chip is devoted to comparative genomic hybridization. It is designed to help clinicians determine the relative amount of a given genetic sequence in a particular patient. "A certain amount of unusual genetic expression is normal, but it becomes a cancer out of control only when the level of expression reaches a dangerous level. As an extreme example, many breast cancer tumors—particularly at the end stages of the disease—are so violently aberrated genomically that they don't even have 23 chromosomes anymore," Müller points out. "This type of chip is designed to look at the level of aberration." This is usually done by using a healthy tissue sample as a reference and comparing it with a sample from the diseased tumor.

Design Goals

In the first installment of this series we reviewed the types of technologies companies are using to manufacture their products. While those basic technologies have not changed, rapid progress is being made in all of them to increase density (the number of arrays per chip) for the next generation of products in development. Whether via robotic deposition or microlithography, development is proceeding in pursuit of two basic design goals: increased sensitivity and reliability, and systems integration.

In the case of the former, the "holy grail is to get better discrimination," says Lance Fors, president of Third Wave Technologies (Madison, WI). "The key to better discrimination is improved signal-to-noise ratio." This is especially important since so many DNA chips depend on PCR-based amplification of the sample, which can undermine discrimination. "A challenge facing us is to find a tumor cell amidst a thousand healthy cells in a biopsied sample; it's like finding a needle in a haystack," he says.

Third Wave and other companies are working to reduce needed sample size by decreasing the amount of reference sample, thus reducing the amount of PCR amplification needed. The reduction of the amounts needed is being accomplished through advances in microfluidic technology. For example, capillary electrophoresis breakthroughs and nanoscale fabrication development have enabled companies to reduce needed reference and target sample sizes to microliter and picoliter scales.

In addition, says Wilding, piezoelectric charges applied to these tiny capillary tubes can move samples from preparation to reaction on a single chip. This achievement has laid the groundwork for the second goal, systems integration. "To do this you must have a microfluidics platform," he says.

"Systems integration is no longer a pipe dream," Wilding continues. "Five years ago an industry survey identified a dozen or so companies developing chips, nearly all of which were involved in detection applications. Three were involved in reactions, and none were doing sample preparation. Today nearly everyone is involved in all of these applications, and all have recognized the importance of an integrated approach."

An example of this work is the collaboration among the U.S. Department of Energy's Argonne National Laboratory, Engelhard Institute in Russia, Motorola, and the Packard Instrument Co. This consortium is working on a microarray-based system that would perform sample preparation (using PCR amplification) and reaction on a single chip and within a single analytical device.

"I think you'll see integrated systems that will address some key markets in clinical chemistry sometime in 1999," predicts Thakkar. He agrees with Wilding that these will be integrated systems designed for ease of use.

Ultimately, within the next 10 years, Wilding and others predict, DNA chips and integrated processing will penetrate the emerging point-of-care market. To achieve this, however, myriad commercialization hurdles must be overcome.

Conclusion

DNA-chip development is proceeding at a pace so rapid that it surprises even the most optimistic members of this fast-emerging industry. "The question 'Will DNA chips succeed?' is a dumb one," notes Wilding. "The only real questions are how they will develop and how quickly."

As companies look to market products outside the research-laboratory and drug-development environments, they will face enormous regulatory and market challenges. Chief among the regulatory challenges is developing a set of industry standards for quality assurance. Today each company has its own. "This will involve some of the big companies like Affymetrix taking the lead, and it will require a real partnership between FDA and all the key players in the industry," says Thakkar.

The biggest marketplace hurdle will be to introduce products at prices that are competitive with existing technologies, says Wilding. "In today's reimbursement climate for health care, products must be better as well as cheaper," he points out. These are issues that will be discussed further in the third and final installment of this series, which will discuss future developments.
 

Cliff Henke is a freelance writer based in Southern California.

Cepheid bridges the sample-volume gap

Kurt Petersen

Regardless of the details of any particular technique, diagnostic applications of DNA-chip technologies must contend with one overriding and frequently overlooked requirement: they must be sensitive to low-concentration target analytes. Modern DNA diagnostic assays face mounting demands to detect organisms or DNA mutations at very low concentrations, often less than 100 copies per ml, in raw biological samples such as blood or urine.

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Magnetic microspheres have been used for some time as the solid phase for immunological tests and assays encompassing a number of formats (see Table I). A major advantage of microspheres over such solid supports as filters, tubes, wells, or large plastic beads, is their ease of separation from an aqueous phase. The magnetic character of the microspheres currently on the market varies, with a popular choice being microspheres that are superparamagnetic, meaning that they retain no magnetic character after being removed from a magnet.

Streptavidin-coated magnetic microspheres. Illustration by Oscar Meza.

 

Recently, several companies have begun to offer these magnetic microspheres preconjugated with some type of generic binding protein. A common choice is streptavidin (from Streptomyces avidinii, or produced recombinantly), which is similar to the avian egg protein avidin. Streptavidin interacts strongly with the molecule biotin, and so by biotinylating the ligand to be coupled to the microspheres, an attachment with a bond strength approaching that of a covalent bond (Ka = 10¹⁵/M) can be achieved in a one-step chemical reaction. This reduces both the time involved in protein conjugation and the expense of wasted reagents.

Table I. Commercial magnetic particle-based assay systems.

An important parameter when choosing streptavidin-coated microspheres is their binding capacity. The general approach to determining the binding capacity of such microspheres is to conjugate a biotinylated ligand that will serve as a sensitive marker. Common markers are detected by chemiluminescence, enzymatic activity, radioactivity, and fluorescence. By knowing the molecular weight of this marker, a reliable estimate can be made regarding the capacity of the streptavidin-coated microspheres to bind a ligand of similar size and weight. Whether developing a binding-capacity assay or relying on the binding capacity reported by the manufacturer, the user should ensure that the reported percent of solids is accurate, since these assays are characterized in terms of the weight of bound ligand per unit weight of microspheres. Therefore, a preliminary gravimetric percent of solids determination is recommended. Finally, the magnetic character of the base particles will play an important role in the ease of handling of the microspheres. Therefore, the efficiency of magnetic separation for various types of base particles should be looked at as well.

Possible Assay Strategies

The fundamental reason for designing a binding-capacity assay for this type of solid support is to simplify subsequent ligand attachment to the microspheres. The idea is to bind a marker of similar size and weight to the ligand that will be used for the final application. Using this number as a guideline for the ultimate coupling reaction removes later guesswork regarding proper reagent usage. This capacity for increased efficiency in reagent usage is one of the reasons that the biotin-streptavidin coupling strategy is becoming more popular than conventional covalent coupling protocols in immunoassay and molecular biology applications.

Table II. Characteristics of different binding capacity assay formats for streptavidin-coated magnetic microspheres.

In developing an appropriate assay strategy, both the means of detection and utility for various sized ligands should be considered. Table II lists four widely used detection methods, as well as common markers for each. The corresponding reaction schemes are illustrated in Figure 1.

Figure 1. Basic binding-capacity assay strategies: (a) large-molecule enzymatic assay using biotinylated alkaline phosphatase with a molecular weight (MW) of approximately 140,000; (b) small-molecule fluorescent assay using biotinylated fluorescein isothiocyanate (MW ~633); (c) small-molecule chemiluminescent assay using biotinylated acridinium (MW ~877); and (d) pure biotin radioactive assay using tritiated biotin (MW ~247).

Four commonly used means of detection, in the order of least to most sensitive, are radioactive (RIA), enzyme-linked (EIA), fluorescent (FIA), and chemiluminescent (CLIA) immunoassays.¹ While immunoassays based on fluorescence are quite sensitive, they can be problematic as well. Although our initial work involved this type of assay, we no longer use it because the equipment necessary for optimal detection (a fluorimeter) is not readily available. Also, difficulty working with FITC-labeled biotin, a common fluorescent tag, made it prohibitive for use as a primary binding-capacity assay. These difficulties included the fact that the FITC-labeled biotin was labile, and that it was difficult to dissolve in an aqueous suspension. For these reasons, this approach is omitted from the following assay descriptions.

Radioactive Assay: Tritiated Biotin

The earliest types of streptavidin binding-capacity assays used ¹²⁵I-, ¹⁴C-, or ³H-labeled biotin as tracers. Because of the simplicity of these assays, they are still widely used today. Radioactivity has some distinct advantages over other means of detection, as it causes only very minor changes to the structure of the labeled antigen (a tritiated biotin has the same molecular size as nonradioactive biotin), is easy to quantify, and is simple to detect.² This makes the use of these materials very convenient for the study of binding reactions of small molecules. Additionally, a large biotinylated molecule can be radioactively labeled to easily determine the binding capacity of streptavidin-coated microspheres for higher molecular weight ligands, such as immunoglobulins. Before deciding to use an apparently simple radioactive assay as the primary biotin binding-capacity assay, however, users should give consideration to such complicating factors as the low specific activity, even of ¹²⁵I-labeled molecules; the labile nature of some radioactively modified molecules; the regulatory pressures and constraints involved in using such tests; and the need for specialized detection equipment.

There are two commonly used approaches to this type of assay. The first is to incubate varying amounts of microspheres with an excess of radioactively labeled biotin to calculate binding capacity. Perhaps the more common approach is to incubate varying amounts of radioactive biotin with a constant amount of microspheres to generate a Scatchard plot. A version of the Scatchard plot is the typical approach for quantifying the number of receptors (i.e., binding sites) on a cell surface, and thus can be modified for quantifiying the number of binding sites on a microsphere surface (see Figure 2). Our approach to this assay was to incubate varying amounts of streptavidin-coated magnetic microspheres with a constant amount of tritiated biotin, always in excess of the stoichiometrically calculated number of biotin binding sites on the microspheres. The microspheres were then washed, and the radioactivity was determined directly from the microspheres by scintillation counting. The actual binding capacity is a conversion of the fraction of counts per minute of bound biotin divided by the counts per minute of total biotin added. For more accurate measurement of tritiated biotin activity, a check can be run to detect the counts per minute of the supernatant, ensuring that this count, plus that for the microspheres, adds up to the counts per minute for the original volume of tritiated biotin.

Figure 2. Example of a Scatchard plot, typically used to quantify the number of receptors on a cell surface.

 

When measuring the binding capacity of our streptavidin-coated magnetic microspheres radioactively, we encountered some interesting considerations. We found that a broader size distribution of base microspheres, such as are offered by several leading suppliers of magnetic particles, can lead to inaccurate and low binding-capacity values. It is believed that this is a result of "fines," or smaller particles that are not pulled to the magnet in the same amount of time as the main population of microspheres. These fines could theoretically bind biotin yet not be detected by scintillation counting of the microsphere pellet, thereby lowering the apparent binding capacity. One way around this source of error would be to use a more powerful magnet with the capacity to pull microspheres that have a lower amount of iron oxide, as is the case with fines.

Enzymatic Activity Assay: Biotinylated Alkaline Phosphatase

As regulatory issues involving radioactive markers have become more prevalent, interest has grown in using other means of detection for binding-capacity assays. A main focus in assay development has been the use of enzyme labels. While several enzymes can be used as markers, one of the most common, because of its widespread commercial availability, is alkaline phosphatase. Reasons for the popularity of this type of binding assay include its simplicity, the strong signal given by the enzymatic reaction with a substrate, and the fact that the size and molecular weight of alkaline phosphatase mirror those of many commonly attached ligands, such as immunoglobulins. By developing a standard curve based on the absorbance of varying concentrations of enzyme per constant concentration of substrate, precise quantitation of binding can be derived simply by colorimetric detection using a spectrophotometer.^3,4

For streptavidin-coated magnetic microspheres, this assay method involves first making serial dilutions of biotinylated alkaline phosphatase (B-ALP) and reacting these with a constant concentration of substrate, in this case paranitrophenyl phosphate (PNPP). The concentration of substrate used will determine the reaction kinetics, so some optimization will be required to find a suitable concentration that allows sufficient time after addition for accurate spectrophotometric measurement. Once this concentration has been determined, the serially diluted B-ALP is reacted with the substrate, and the absorbance readings at 405 nm are used to generate a standard curve. The proper dilutions for the B-ALP are determined such that they will fall within the range of linearity for absorbance measurements on the spectrophotometer that is used.

Once these variables have been optimized, the last variable to consider is the concentration of microspheres to be used in the actual assay. As the B-ALP conjugated microsphere concentration increases, so does the amount of signal given off by the ALP-PNPP interaction. Therefore, a microsphere concentration must be established such that the signal is within the limits established by the standard curve.

An important precaution with this assay is the time allowed for substrate development (color formation). As ALP is allowed to react with PNPP, color continues to form until all of the substrate has been exhausted, giving the potential for a false high reading. Therefore, if a rate-dependent format is chosen, the time allowed for color development must be precisely controlled. Conversely, if an end-point reading is to be taken, the relative concentration of the ALP-conjugated microspheres must be controlled, such that allowing the reaction to go to completion will still give absorbance readings within the limits of the standard curve.

Chemiluminescent Assay: Biotinylated Acridinium

Chemiluminescence is the chemical generation of visible light by a reaction, and as such does not use any light source. Thus the need for complicated and inefficient optical wavelength filtering systems is eliminated. Chemiluminescent systems fall into two classes. The first and easiest to develop uses enzymes to produce the chemiluminescent signal. Typically, either horseradish peroxidase or alkaline phosphatase is used, and the label is triggered by the addition of substrates that, under the influence of the enzyme system, give rise to a visible emission. This type of signal enhancement has enabled researchers to develop binding-capacity assays that are faster and more sensitive than any traditional colorimetric or radioactive assay.

The other chemiluminescent systems use a nonenzymatic direct chemiluminescent label. Direct labels tend to have lower background signals than enzyme systems, and typically produce a signal very quickly. With the acridinium ester system, after the immunological binding and subsequent wash step, the signal takes only 2 seconds to develop, compared with 30 minutes or longer for an enzyme-generated system (see Figure 3).

Figure 3. Acridinium C^₂HNS ester (formula weight 632.55) can be biotinylated and used to develop a rapid and sensitive small-molecule biotin-binding assay.

 

As luminometers used to detect chemiluminescence have become more common in today's laboratories, the interest in chemiluminescent binding assays for the quantitation of streptavidin-coated magnetic microspheres has increased. The advantage that chemiluminescence offers over colorimetric enzyme, fluorescent, or radioactive assays is enhanced sensitivity.⁵ Of particular benefit when developing a binding-capacity assay specific to streptavidin-coated magnetic particles is that chemiluminescent and radioactive detection are the only formats that can be read in the presence of the microspheres without interference from the particles themselves.

This newer approach to developing a binding-capacity assay carries a disadvantage. Unlike the more conventional EIA, RIA, and FIA approaches, the commercial availability of well-characterized chemiluminescent esters that are able to be biotinylated is limited. Were this not the case, a direct binding-capacity assay could be run by incubating the chemiluminescent ester with the microspheres, measuring with a luminometer the relative light units (RLUs) emitted by a certain concentration of microspheres, and then converting this to actual binding based on the number of RLUs given off by one chemiluminescent molecule. As it is, a chemiluminescent ester can be biotinylated and purified. However, unless the concentration of acridinium in a given sample is known precisely, it is necessary to develop the assay using an indirect format.

With this obstacle in mind, such an assay was carried out by first flooding the microspheres with free biotin, washing, adding a constant concentration of biotinylated acridinium (B-ACR), and rewashing (see Figure 4). This ensured that, based on stoichiometry derived from the titration for the number of groups on the base particle available for streptavidin binding, all of the binding sites on the streptavidin were occupied by free biotin. This was used as the "blank" on the luminometer, and any signal given off was attributed to nonspecific binding of the acridinium ester (a hydrophobic molecule) to the exposed surface of the hydrophobic microspheres. Optimization of the blockers used in the assay or more-vigorous washing steps are necessary to lower this value to near zero.

Figure 4. Increasing chemiluminescent signal, as read on a luminometer, results when a lower concentration of free biotin is added in the presence of a constant amount of biotinylated acridinium.

 

We then serially diluted the free biotin, and incubated this with the same constant concentration of B-ACR used previously, and streptavidin-coated magnetic microspheres. As the luminescent signal increased from the washed particles, this was an indication of the number of free sites left open for binding of the biotinylated acridinium. The dilutions were carried out to complete a sigmoidal binding curve, with the dilutions representing the steepest part of the curve being narrowed for increased precision (see Figure 4). Finally, as it was difficult to identify a direct point on the curve to assess maximum binding, we reasoned that the point on the curve with the steepest slope could be considered to be the equivalence point (with half of the streptavidin binding sites occupied by biotin, and half by B-ACR), and that multiplying this value by two would give a precise binding capacity value for B-ACR.

Factors Influencing Assay Variability

Although the total binding capacity of streptavidin-coated magnetic microspheres is an important factor for most applications, with the highest total binding normally being the best, other variables play important roles. Because binding capacity measurements are given in terms of a weight of bound marker per unit weight of microspheres, the percent of solids in the suspension in which these are supplied is important for obtaining their true binding capacity. Before designing any assay, we recommend measuring this percentage gravimetrically to ensure that the measured percent of solids coincides with the reported percent of solids. Figure 5 illustrates differences that we found in our lab for material supplied by several leading vendors.

Figure 5. Variables involved in determining the binding capacity of streptavidin-coated magnetic microspheres: (a) relative total binding and nonspecific binding capacity of microspheres from several suppliers based on supplier-reported percent of solids (R) and on experimentally determined (gravimetric) percent of solids (EX); (b) effect of adding and optimizing a blocker to reduce nonspecific binding of the acridinium label.

 

Similarly, the total binding can be classified in terms of both specific and nonspecific binding. The former refers to biotinylated ligands that are attached via the streptavidin-biotin bond, whereas the latter refers to ligands that are attached via some other mechanism—normally hydrophobic adsorption to the particle surface (see Figure 5). By designing the binding-capacity assay so that both types of binding can be measured, optimization using various types of blocking molecules can be performed to minimize nonspecific binding.

Conclusion

In the fields of immunology and molecular biology, streptavidin-coated magnetic microspheres offer several advantages over more-conventional solid supports. But to use this type of solid support to the fullest, it is important to first ensure that the binding capacity is fully characterized for the type (size and molecular weight) of ligand that the final application will ultimately use.

Expectedly, as the technology involved in making this type of solid support has advanced, so has the sensitivity and simplicity of the assays used to characterize them. As immunoassays have evolved from radioactive to enzymatic to chemiluminescent detection, binding-capacity assays have followed.

The point that we have tried to stress in this article is that once the choice has been made to base an immunoassay on streptavidin-coated magnetic microspheres, it is important to select the microspheres based on their performance in the assay system under development. Variables that influence performance are not just total binding, but binding of molecules with steric characteristics similar to those of the molecules that will be used in the actual assay. If this information is not available from the manufacturer, one of the assays described here can be used for appropriate characterization. Further, among the streptavidin-coated magnetic microspheres currently on the market, the characteristics of the base particles vary greatly. In developing these binding-capacity assays, we looked at particles from a number of suppliers, and made comparisons based not only on total binding, but on characteristics such as nonspecific binding, ease of handling (how rapidly they could be pulled to a standard magnet), measured versus reported percent of solids, and so on. We feel that these considerations are vital in ensuring that the reagent selected is truly the best choice for a particular application.

References

1. Hart R, personal communication, Ann Arbor, MI, Assay Designs, Inc., (www.assaydesigns.com), 1998.

2. Price CP, and Newman DJ, Principles and Practice of Immunoassay, New York City, Stockton Press, 1997.

3. Harlow E, and Lane D, Antibodies: A Laboratory Manual, Cold Springs Harbor, NY, Cold Springs Harbor Laboratory, 1988.

4. Savage D, Mattson G, Desai S, et al., Avidin-Biotin Chemistry: A Handbook, Rockford, IL, Pierce Chemical, 1992.

5. Wild D, The Immunoassay Handbook, New York City, Stockton Press, 1994.

Joseph M. Duffy, John V. Wall, and Mary B. Meza are members of the technical staff at Bangs Laboratories (Fishers, IN). Laura J. Jenski, PhD, is a professor of biology at Indiana University–Purdue University (Indianapolis).

Over the years, very few constitutional challenges to FDA authority have been successful. But on July 30, Judge Royce Lamberth of the United States District Court for the District of Columbia struck down several important FDA policies on the ground that they violated the First Amendment.¹ This decision may benefit IVD companies.

Historically, FDA has prohibited companies from distributing information about off-label uses of their products except in certain narrowly defined circumstances. FDA significantly restricted the dissemination of scientific articles that discuss off-label uses of approved drugs, biologics, and devices. The agency also developed policies curbing company-sponsored seminars at which off-label uses were discussed.²

The WLF Suit

In 1994, the Washington Legal Foundation (WLF; Washington, DC) filed a lawsuit challenging FDA's restrictions on the dissemination of information relating to off-label uses. WLF's principal allegation was that FDA's policies infringed upon the First Amendment's protection of free speech.

In response, FDA sought to have the case dismissed on procedural grounds. For example, FDA asserted that since the restrictions were merely statements of policy—as distinguished from rules—there was no final agency action reviewable by the court. The agency also argued that WLF did not have standing to bring the case because it was not directly affected by FDA's restrictions. Judge Lamberth rejected both of these arguments.³

Although most lawsuits involving FDA are decided without the plaintiff being permitted to depose any agency employees, the discovery phase of this case presented an exception. WLF was allowed to take depositions from several senior FDA officials, including William Hubbard, associate commissioner for policy coordination, and Byron Tart of the Center for Devices and Radiological Health. The court found some of the information gleaned during these depositions to be relevant to its decision.

The discovery process also resulted in an unprecedented ruling. Over FDA's objections, WLF was granted the right to depose then-commissioner David Kessler. FDA sought relief in the court of appeals, which was denied.⁴ If not for his subsequent resignation, Kessler would have faced the dilemma of whether to allow himself to be deposed or potentially be found in contempt for refusing to be deposed.

The July Decision

After all this procedural wrangling, both FDA and WLF filed motions for summary judgment. In its July 30 order, the court denied FDA's motion and granted WLF's.

The first issue that the court needed to address was whether FDA's policies regulated speech or conduct. This threshold issue was critical, because conduct is not entitled to the same level of constitutional protection as speech. The judge disposed of FDA's argument quickly, saying that "This court is hard pressed to believe that the agency is seriously contending that 'promotion' of an activity is conduct and not speech, or that 'promotion' is entitled to no First Amendment protection."⁵

FDA next argued that First Amendment protections did not apply because the agency has pervasive powers to regulate industry, in effect subsuming industry's First Amendment rights. The court disagreed, ruling that government's power to regulate many industry activities does not mean that it also has an untrammeled right to regulate speech.

However, the court agreed with FDA on the next point: that the distribution of reprints and sponsorship of scientific symposia constitute commercial speech. WLF had argued that these forms of communication are pure speech, which is entitled to a much higher level of constitutional protection than commercial speech. Governmental restraints on pure speech rarely survive judicial scrutiny.

In finding that these activities are commercial speech, the court used language that is likely to be cited by FDA in the future. Referring to the possible risks of enduring materials and company-sponsored seminars, the court wrote, "The potential to mislead, and the harm that could result, convinces this court that it is permissible to 'depart from the rigorous review that the First Amendment generally demands.' "⁶

Having decided that reprints and seminars constitute commercial speech, the court then applied the four-prong test established by the Supreme Court for reviewing the constitutionality of restrictions on commercial speech. First, the court found that the speech was neither unlawful nor inherently misleading. The agency had argued that until FDA has reviewed and approved a claim, it is inherently misleading. In a passage that is likely to be frequently quoted, Judge Lamberth wrote:

In asserting that any and all scientific claims about the safety, effectiveness, contraindication, side effects, and the like regarding prescription drugs are presumptively untruthful or misleading until the FDA has had the opportunity to evaluate them, FDA exaggerates its overall place in the universe. . . .[The] conclusions reached by a laboratory scientist or university academic and presented in a peer-reviewed journal or textbook, or the findings presented by a physician at a CME seminar, are not "untruthful" or "inherently misleading" merely because the FDA has not yet had the opportunity to evaluate the claim.⁷

Nevertheless, the district court also emphasized that its ruling does not restrict FDA's ability to control claims that actually are false or misleading. In determining what actions to take in light of the WLF case, companies will need to pay heed to this important limitation on the scope of the ruling.

Applying the second element of the Supreme Court's test, the court agreed that FDA has a substantial interest in regulating enduring materials and seminars, and in forcing manufacturers to obtain agency approval for off-label treatments. (Government regulation of commercial speech can be voided if such regulation does not advance a substantial interest.) But the court also said that FDA cannot justify its regulation by citing fears that such commercial information might be misused by physicians.

The court further agreed that FDA's policies advance this important interest. The court observed that FDA's restrictions on enduring materials and continuing medical education (CME) programs provide an incentive for manufacturers to get approval for off-label uses of their products.

While agreeing that FDA's restrictions advanced a substantial government interest, however, the court found that they flunked the fourth prong of the Supreme Court's test: they are more extensive than necessary. The court advanced several reasons for its conclusion. For instance, it noted that off-label treatments are often the most effective ones. "In this case, the truthful information may be life saving information, or information tha[t] makes a life with a debilitating condition more comfortable."⁸ The court also noted that even without these restrictions, companies have adequate incentives to get FDA approval for new uses.

More Wrangling

Even the court's July decision has not brought an end to the procedural wrangling over this suit. Since the decision was handed down, FDA has filed a motion to amend the judgment, basing its argument on the extensive provisions relating to the distribution of off-label reprints and textbooks in the FDA Modernization Act of 1997 (FDAMA).⁹ The provisions of FDAMA would allow such distribution, but only under circumscribed conditions and with FDA's prior approval. FDA has asked that the court's order be modified so that it would affect only the agency's guidance documents, and not FDAMA. WLF has opposed that motion, arguing that the court's First Amendment decision applies to all FDA restrictions on the dissemination of speech—including those in FDAMA.

FDA's motion is currently pending. If it is granted, FDA's current guidance documents on this subject will be void, but companies will be required to comply with FDAMA's restrictions on the distribution of reprints when that section goes into effect this November. CME programs, which are not mentioned in FDAMA, will be subject only to the court's order, and not to FDA's guidance documents. If the motion is denied, the court order issued on July 30 would presumably prohibit the agency from using FDAMA to restrict the distribution of reprints or textbooks relating to off-label uses.

In industry, there has been some misunderstanding of the meaning of the court's order. It certainly does not mean that companies can distribute any off-label information they want. However, under the order FDA cannot prohibit an IVD company from taking the following actions:

• Distributing reprints of articles that have appeared in a bona fide, peer-reviewed professional journal. FDA could interpret this provision as allowing it to restrict the distribution of special supplements devoted to a single product.

   

• Distributing textbooks published by a bona fide independent publisher; that is, a publisher that has no corporate affiliation with the manufacturer and is in the business of publishing and distributing books.

   

• Suggesting content or speakers for an independent program provider. A provider is independent if it has no connection with the sponsor, engages in the business of creating CME seminars, and is accredited by a national organization.

Significantly, the order explicitly allows FDA to take action to prevent the distribution of materials that are false or misleading. Moreover, FDA can require companies to disclose that the uses described in the article or discussed at the seminar have not been approved by the agency, and can compel the company to disclose its interest in the drugs or devices being discussed. IVD companies would be well advised to take these steps, even in the absence of an FDA requirement.

IVD Implications

Although the WLF case concerned both devices and drugs, the court's opinion does not specifically refer to IVDs. Nevertheless, IVD companies can benefit from the decision if they understand what that decision means to them.

Assuming that FDA's motion to amend the judgment is not granted, an injunction is in effect prohibiting FDA from taking enforcement action against informational activities that were previously prohibited. However, companies that decide to distribute reprints should read the court's order carefully, and comply with all of its limits. They should also be judicious in which articles they choose to distribute.

There are certainly numerous instances of IVDs with off-label uses for which a substantial literature exists. One example is the use of alpha-fetoprotein (AFP) testing as a marker for Down's syndrome. Although AFP is often used for this purpose, such a use has not been cleared by FDA. Nor has the widely used triple-marker screen been approved by FDA. Companies will be able to distribute reprints relating to these uses if they are within the scope of the court's order.

Similarly, prostate-specific antigen (PSA) assays are routinely used to screen for prostate cancer, even though they are not approved for that purpose. The court's ruling would potentially permit companies to distribute certain articles regarding this off-label use. Of course, this would not provide equal footing with an assay approved for screening; an approved product can be advertised for screening, while an unapproved assay cannot. Even so, the ruling opens up new opportunities to provide information about off-label uses.

FDA has also restricted the flow of information regarding research-use-only and investigational-use-only products.

The court's ruling does not directly address these categories of IVDs. Nevertheless, the WLF case raises potentially significant constitutional issues for FDA policies that restrict the advertising of these products.

Conclusion

The WLF lawsuit has already lasted four years, and it may stretch on even longer. FDA may well choose to appeal the court's decision. How any appeal will come out is anyone's guess. The court of appeals could affirm the decision, reverse it on the procedural grounds raised by FDA long ago, or reverse it on the merits of the case.

Ironically, perhaps FDA can take comfort from its recent loss in the tobacco case. That case illustrates that a district court victory in a high-profile FDA case doesn't always mean success in the court of appeals.

References

1. Washington Legal Foundation v. Friedman, Civ. No. 94CV01306 (U.S. District Court, District of Columbia [DDC], July 30, 1998).

2. Gibbs J, "Industry-Supported Educational Programs: FDA's Final Policy," Regulatory Affairs Focus, 3(3):22, 1998.

3. Washington Legal Foundation v. Friedman, 880 F. Supp. 26 (DDC, March 9, 1995).

4. Washington Legal Foundation v. Friedman, 100 F.3d 1015 (D. C. Cir., November 29, 1996).

5. Washington Legal Foundation v. Friedman, Civ. No. 94CV01306, p 16.

6. Washington Legal Foundation v. Friedman, Civ. No. 94CV01306, p 32.

7. Washington Legal Foundation v. Friedman, Civ. No. 94CV01306, p 36.

8. Washington Legal Foundation v. Friedman, Civ. No. 94CV01306, p 53.

9. Food and Drug Administration Modernization Act of 1997, Pub. L. 105–115, sec. 552.

Jeffrey N. Gibbs is a partner in the law firm of Hyman, Phelps & McNamara (Washington, DC).

Perceptive Scientific Instruments (PSI; League City, TX), a subsidiary of International Remote Imaging Systems (IRIS; Chatsworth, CA), has been awarded a grant for a study of how to deblur fluorescent microscope images. The phase I small business innovative research grant was awarded by the National Center for Research Resources and will be used to determine the feasibility of new methods to improve microscope images.

To deblur FISH images, PSI is studying the use of software that can compose a single image (bottom) from images of two separate microscopic planes (top, center). Photos Courtesy PSI, Inc.


 

With any type of microscopy, focusing on structures in two different planes is difficult. As one structure comes into focus, the other becomes blurred. With automated microscope screeners and slide-reading systems, the problem becomes even greater. With this study, PSI is focusing its research on ways to eliminate this problem when analyzing specimens prepared by fluorescent in situ hybridization (FISH).

According to PSI senior research engineer Mark Schulze, "one of the approaches will be to take images from two different planes and then use software to fuse them into a single composite image." Another strategy might involve deconvolution of a single image to bring all structures back into focus, he adds.

If successful, the technology figures to strengthen PSI's Powergene line of products. Powergene products are currently used for cytogenetic procedures, including automatic karyotyping and FISH chromosome painting for comparative genomic hybridization analyses.

IRIS manufactures and markets automated in vitro diagnostic systems for clinical and research laboratory procedures.—G.W.

In June, Johnson & Johnson subsidiary Lifescan, Inc. (Milpitas, CA), announced a product recall for its Surestep blood glucose meters. According to the company, units made before August 1997 can report an "Er 1" error message when blood glucose levels are higher than 500 mg/dl. Unless they receive treatment, patients with glucose levels in that range can experience serious medical consequences.

Although Lifescan initially characterized the recall as a product replacement program, FDA classified the company's action as a Class I recall, meaning that a reasonable probability exists that continued use of the product will cause serious adverse health consequences or death. FDA has received reports of two deaths of diabetics who delayed seeking medical care after their Surestep meters gave an error message.

Nevertheless, FDA officials recommended that diabetics continue using their Surestep monitors to measure their glucose levels, with the caveat that an "Er 1" message can mean a high glucose level. They also stressed that it is far more dangerous for diabetics not to check their glucose levels than to continue using a glucose meter that could give unclear messages.

Like the Lifescan monitor, most other digital glucose monitors use electronic algorithms to translate and display their data. According to FDA, however, the Surestep is the only device reported to have problems of this sort.

Along with its digital readout, the Surestep also provides colorimetric data on its test strips. If the digital display provides any ambiguous results, consumers are being urged to compare the test-strip indicator to the color chart printed on the test-strip packaging. An indicator dot that is as dark as or darker than the color chart indicates a very high glucose reading, and the user should seek treatment.

Lifescan is replacing faulty units free of charge and has set up a hotline (800/951-7226) to educate consumers about the monitors.—G.W.

Xanthon, Inc. (Research Triangle Park, NC), was recently awarded a $200,000 economic development loan from the North Carolina Biotechnology Center to support development of the company's electrometric technology for the detection of DNA and RNA.

Using this financing, together with a credit line of $500,000 recently secured from Imperial Bank of California, Xanthon expects to continue development toward commercialization of its product in 18 to 24 months.

Xanthon's electrometric system is a platform for molecular assays based on the electrochemical detection of DNA or RNA. When a sequence-specific probe hybridizes, it creates an electric current that can be measured and reported by the system.

A low-cost alternative to current platforms for molecular diagnostics, Xanthon's electrometric system detects electric current produced when a sequence-specific probe is hybridized.

 

Promotion of the company's nucleic acid detection technology will be aimed primarily at the market for high-throughput drug discovery, but also at companies involved in the development of molecular diagnostics for infectious diseases and cancers. According to Xanthon president and CEO James D. Skinner, the company's system "can be used anywhere that expensive optical detection methods, such as fluorescence, are used."

Suggesting that Xanthon's technology will provide a low-cost alternative to current platforms for molecular diagnostics, Skinner says that one of the major reasons for the slow adoption of molecular assays is their prohibitive cost. "Pivotal to the widespread adoption of molecular assays will be the ability to make them cheaper."—G.W.

Medical product engineers, designers, manufacturers, and distributors are invited to submit their products to the 1999 Medical Design Excellence Awards, a program that recognizes innovative medical devices. The competition is sponsored by Canon Communications llc (Santa Monica, CA) and endorsed by the Industrial Designers Society of America (IDSA; Great Falls, VA).

Bruce Behringer (left) and Greg Farrell of Bayer Diagnostics (Tarrytown, NY) accepted the 1998 MDEA award for the Unifluidics module, an integral part of the company's Advia 129 hematology blood analyzer.

 

An eight-member jury will evaluate the submissions for design and engineering innovation, contribution to health care, cost-effectiveness, benefits to users and patients, and business impact. The jurors are Sohrab Vossoughi, principal of Ziba Design (Portland, OR); Kent Ritzel, a director of Metaphase (St. Louis); John Gosbee, MD, director of the Center for Applied Medical Informatics, Michigan State University Kalamazoo Center for Medical Studies; Matt Duncan, president of Morphix Design (San Clemente, CA); Christoph Böninger, executive board member, Siemens Design & Messe GmbH (Munich, Germany); Robert Hall, principal, GVO (Palo Alto, CA); Michael Wiklund, director and principal research scientist, American Institutes for Research New England Research Center (Concord, MA); and Corinna Lathan, assistant professor of biomedical engineering at the Catholic University of America (Washington, DC).

To be eligible for submission, products must be commercially available by the February 8, 1999, deadline. Competition rules and entry forms are available by calling the 24-hour Fax-on-Demand line at 800/588-8527, by visiting Medical Device Link at http://www.devicelink.com/awards, or by calling Canon Communications at 310/392-5509 or IDSA at 703/759-0100.

Winning products will be on display throughout the Medical Design & Manufacturing East 99 Conference and Exposition, which will be held in New York City, May 25–27, 1999.

Makers of integrated testing systems may be encouraged to know that demand for their equipment among laboratorians is remaining steady. But companies producing total laboratory automation equipment may have a long way to go before they convince potential purchasers to buy their systems.

Those are among the key results of a survey of senior-level lab managers conducted by the Clinical Laboratory Management Association (CLMA; Wayne, PA) last May and June, and released in October. Responses from the 495 managers who participated in the survey suggest that robotics and automation will be adopted by a greater number of clinical laboratories in the coming year.

Only 7.7% of the responding managers said they had implemented robotics or automation in the past year, but 16.8% said they expected to do so in the coming year. The forms of automation that labs are most likely to implement over the next five years include integrated testing systems (41%), automated centrifugation (37%), and automated aliquoting (35%), the survey found (see Table). Total laboratory automation was far down the list at 12%. The percentages were virtually identical to a CLMA survey conducted the previous year, suggesting that demand for automated testing equipment is likely to remain stable over the next five years.

Percent of respondents that expect named types of laboratory automation to be implemented in their clinical laboratories within the next five years, according to surveys conducted in 1997 and 1998 by the Clinical Laboratory Management Association.

Technology purchases are only one way that laboratory managers are seeking to streamline performance and reduce turnaround times. The survey results suggest that significant restructuring is continuing among clinical laboratories.

One result of this restructuring seems likely to be the demise of self-standing and independent clinical laboratories. More than a third of the respondents (37.4%) said that their laboratory had joined a network or alliance in the past 12 months, and nearly a third (29.3%) indicated that they would be doing so in the next 12 months.

Another restructuring strategy that appears to be on the rise is the concept of implementing a core lab: 18.8% of respondents did so in the past 12 months, and 22.0% expect to do so in the next 12 months. Managers indicated it is common for them to use a combination of strategies.

Some restructuring strategies, however, appear to be on the wane. More than a third of respondents (35.8%) indicated that they had adopted a point-of-care testing program in the recent past, but only 9.9% expect to do so in the near future. Outsourcing of tests has already been adopted by 9.7% of labs in the past, but is expected in the future by only 8.5% of respondents. Best of all for laboratorians, downsizing, which has already been carried out by 30.5% of labs, is expected by only 17.8% in the future.

The survey results also indicate some changes in the types of tests that laboratories are choosing to keep in-house or outsource. Among the tests most commonly brought into the labs during the past year, thyroid tests were the most common (brought in by 23% of respondents' labs). Among the tests outsourced, molecular/PCR tests were the most likely to have been sent elsewhere (by 23% of labs), but this trend was nearly balanced by the high percentage of labs (17%) that added molecular/PCR tests to their repertoire. Electron microscopy was also a common outsourcing target (18% of labs reported doing so).

The managers who saw increased point-of-care testing reported an average increase of 10%; but this was balanced by those who saw a decline, also reported at 10% on average. Those who increased outsourcing did so by 5%; those who decreased it did so by 10%.

For about a third of the responding companies, all of these technology purchases and restructuring strategies appear to be paying off in quicker turnaround times for testing. About 33% of respondents said their facilities showed a drop in turnaround times during the previous year. Much of that drop seems to have come from reduced testing time, where 34% of respondents saw a decrease, compared to the 21% of respondents that reported a decrease in pretesting time and the 21% that saw reductions in posttesting time. For 14% of the respondents' companies, turnaround times increased during the previous year; 52% reported no change.

To purchase a copy of the report, contact Collette A. Steward, PhD, director of research, CLMA, at 610/995-9850, ext. 236.

For the tidy sum of $1.1 billion, Bayer Group is now the proud owner of Chiron's IVD business. By acquiring Chiron Diagnostic Corp. (Walpole, MA), Bayer Diagnostics (Tarrytown, NJ) will expand its product line and developmental pipeline to encompass technologies in molecular diagnostics, immunodiagnostics, and critical-care blood gas analyzers.

Bayer will now have a solid platform from which to enter the rapidly growing field of molecular diagnostics. Mark Ryan of Bayer's media relations department says that, "the technology of nucleic acid diagnostics was the prime reason for the acquisition." Although the acquisition has been announced, the final and official agreement will not be reached for several weeks.

The decision of Chiron Corp. (Emeryville, CA) to sell off its diagnostics business was, in part, a bow to the realities of the competitive marketplace. "To be a strong competitor in an IVD industry that is seeing increasing consolidation, a company needs a certain critical mass, which Chiron Diagnostics didn't have," says Julie Wood of Chiron's corporate communications department. "Chiron Corp. felt it could more effectively participate by divesting certain aspects of its IVD business to Bayer, while retaining some intellectual property rights and enjoying the royalties from them."

Rights that Chiron Corp. will retain include those for its hepatitis C and human immunodeficiency virus blood-screening assays. Chiron will continue its blood-screening business, but will invest more of its resources into areas of therapeutics and vaccines. According to Chiron Diagnostics' corporate communications manager, Judith Rossi, Chiron Diagnostics is no longer an entity and the division is expected to be under full Bayer management within four months.

Over the next few months, diagnostics manufacturers will probably be hearing a great deal about design control. And depending on how well they are already complying with that portion of FDA's quality system regulation (QSR), that could be either good or bad news.

As many in industry will recall, FDA's desire to create requirements for design control was one of the motivating elements that led the agency to turn its 1978 good manufacturing practices (GMP) regulation into the 1996 quality system regulation. As embodied in the QSR, the agency's design control requirements follow a pattern already established by ISO 9001, the fullest of the quality systems standards compiled by the International Organization for Standardization. Device manufacturers were to have completed implementation of their design control systems by last June, after a one-year transition period.

From very early in the process of transforming the GMP regulation into the QSR, diagnostics manufacturers asserted that the application of design controls to the creation of IVDs is very different from their use in other sectors of the device industry. And for just as long, they have been asking for a design control guidance specific to IVD manufacturing. These requests were acknowledged last year by Steven Gutman, director of FDA's Division of Clinical Laboratory Devices, when he identified such a guidance as one of his division's priorities for 1998.

The actual work of compiling such a guidance, however, fell to industry. More specifically, that work has been taken up by a subcommittee of the Association of Medical Diagnostics Manufacturers (AMDM) led by Robert James. According to James, a draft of the committee's work is now being circulated for review, and he hopes that it can be readied for publication by the beginning of next year.

Such a schedule could make the guidance a very timely tool, because FDA's Los Angeles District Office is now working with diagnostics manufacturers to develop a January workshop focused on design control and process validation for the IVD industry. Observers indicate that agency inspections have revealed some reason for concern over industry's compliance with design control requirements. Topping the list of Form 483 observations made by the office during 1998 are erroneous or absent design control procedures. As this issue goes to press, details of the planned meeting are sketchy, but updates will be available via the Web site of the Orange County Regulatory Affairs group, at http://www.ocra-dg.org.

The concerns to be addressed by the January meeting are serious ones that could have a forceful impact on the ability of diagnostics manufacturers to do business. Now that the design control requirements are fully and formally implemented, companies that haven't already complied with them won't be given much leeway when it comes time for an FDA inspection.

But companies shouldn't feel that they need to work out the problems of implementing design control by themselves, either. Fortunately, there are a number of IVD-related organizations that can help, as indicated by this year's directory of organizations and associations. Keeping in touch with others in industry is an excellent way for manufacturers to bring themselves up to date on the latest issues, and to avoid falling short in important areas. This year's directory offers a starting point, but it will be up to manufacturers to make the most of their contacts.