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Companies, Researchers Tout Integrated Microfluidic Platforms at Sample Prep Meeting

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By Ben Butkus

At the Knowledge Foundation's Sample Prep 2010 meeting, held last week in Baltimore, Md., microfluidics, integration, and automation were oft-cited as future drivers of the sample prep space.

The theme of the conference was sample preparation for virus, toxin, and pathogen detection, and a number of executives and scientists from small companies and academic laboratories presented technologies that aim to miniaturize and automate the sample prep process and integrate it on a single platform with downstream amplification and detection technologies.

As such, the overwhelming majority of the presented technologies are small benchtop or handheld devices taking aim at the point-of-care diagnostic or environmental sampling markets.

PCR Insider recently reported on a pair of companies that presented at Sample Prep 2010: Abbott subsidiary Ibis Biosciences, which is currently beta testing an integrated microfluidic sample prep card and instrument that lyses cells, extracts nucleic acids, and performs whole-genome amplification for downstream whole-genome sequence analysis; and Ithaca, NY-based startup Rheonix, which last month netted $12.6 million in financing to support development of its polymeric, integrated sample preparation, nucleic acid amplification, and endpoint detection card.

Following are brief summaries of some of the other notable technologies presented at the conference.

Lawrence Livermore

Raymond Mariella, a senior scientist at Lawrence Livermore National Laboratory, described his lab's efforts to isolate viruses from clinical samples, specifically nasophayngeal swabs, in an effort to combat emerging pathogens such as West Nile virus and the SARS coronavirus.

Currently, Mariella said, the "state of the art" in sample prep for virus discovery and viral genomics uses a combination of physical filters and CsCl-gradient ultracentrifugation. This process, while effective, is labor- and time-intensive.

Picking up on some previous work conducted at the University of California, San Diego, Mariella and colleagues are instead developing a filter system that uses a phenomenon called dielectrophoresis, in which a non-uniform electric field is used to exert force on and move a dielectric particle.

Mariella said the goal of their project is to develop a system that can process environmental or clinical samples in an automated fashion using a modular, small instrument. More specifically, he said that they want to be able to process a milliliter of sample in 10 minutes or less using a microfluidics system that processes only microliters of sample at any given time.

Their technology, which Mariella said is "simple to fabricate," uses a negative-force dielectrophoretic filter to isolate virus particles from other undesirable particles such as bacteria and contaminants. Then, using a variation of electrophoresis called isotachophoresis, they are able to separate exogenous nucleic acids out of a microfluidic stream containing virus particles of interest.

Once these particles are isolated, the group lyses them and performs downstream genomic analyses using techniques such as digital PCR, microarrays, or sequencing.

In particular, Mariella said that his group has been working with RainDance Technologies to incorporate its digital PCR technology into an assay that would be performed following the LLNL-developed sample prep method.

Atlas Genetics

Meantime, John Clarkson, CEO of UK-based diagnostic shop Atlas Genetics, presented his company's technology, called the Atlas Velox, a point-of-care sample prep and diagnostic instrument that incorporates a microfluidic card with a portable, slightly bigger than handheld readout instrument.

"Sample prep is often the most challenging part of point-of-care diagnostics," Clarkson said during his presentation. Another challenge in developing POC diagnostics is that "it has to integrate a wide range of technologies," which means that companies often must license or develop a lot of intellectual property.

Atlas' Velox platform is built on "novel, patented technology," Clarkson said. It includes a disposable, integrated microfluidic cartridge that integrates sample processing, PCR amplification, and labeling of target DNA with an electrochemical probe.

The cartridge is a "stack" of injection-molded parts, the bulk of which is taken up by sample prep, which "fluidically is the most difficult part," Clarkson said.

The cartridge is inserted into the Velox readout instrument, which detects the electrochemical probe-labeled DNA using a technique that is similar to the electrochemical detection method used in blood glucose monitoring.

This detection technology is "cheap and easy," Clarkson said, and eliminates many of the concerns that go along with optical detection methods such as channel crosstalk and artifacts.

Overall, the system can complete a diagnostic test in about 20 minutes; can perform multiple molecular and immuno-based tests alike from a single patient sample; and is portable and inexpensive, with a reader that is manufactured for less than $2,000 and consumables costs of less than $4.

Atlas is initially preparing Velox for the sexually transmitted disease market, including diseases such as chlamydia, gonorrhea, syphilis, and HIV. Thus far it has already tested the clinical sensitivity and specificity of a chlamydia test in collaboration with the John Radcliffe Hospital in the UK, and found that the assay performed favorably in comparison to the established ProbeTec ET test offered by Becton Dickinson.

Atlas expects its first chlamydia test to launch in Europe in 2011.

In addition, the company may tackle the veterinary market by developing POC diagnostic tests for the "strangles," one of the most frequently diagnosed equine infectious diseases, caused by the bacterium Streptococcus equi.

Clarkson said that the company's efforts in this area are being supported by funding from the Wellcome Trust.

Boreal Genomics

Also at the Sample Prep conference, Andrea Marziali, president and chief technology officer of Vancouver, BC, startup Boreal Genomics, presented and provided an update on the company's Aurora platform for nucleic acid sample prep with capacity for integrated sequence enrichment.

The Aurora, Boreal's most recent iteration of a platform that has been under development for several years, uses a technology called synchronous coefficient of drag alteration, or SCODA.

First covered in early 2009 in PCR Insider sister newsletter In Sequence, SCODA uses rotating electric fields to selectively concentrate nucleic acids in a "focusing" gel and separate them from difficult contaminants such as particulate matter and PCR inhibitors (In Sequence, 1/27/2009).

More specifically, SCODA selects molecules based on characteristics such as charge species, linear charge density, size, and stiffness or conformational entropy. The company has had previous iterations of the technology in early-access labs for more than a year, and has demonstrated that it has100-fold-higher yield and purity than standard elution columns when dealing with difficult samples.

Boreal is targeting a wide market for its instrument, and has previously demonstrated the platform's ability to extract DNA from difficult forensic samples including blood on denim and soil; from heavy tar sands in Alberta, Canada, in a proof-of-principle experiment that was published last year in PNAS; from 60-year-old plant leaves; and from clinical samples such as blood and urine.

But one of the latest improvements to the instrument, Marziali said, is that the SCODA technology can be made specific to the sequence of DNA targets to be concentrated, which opens the door to the sequence enrichment market for Boreal.

To achieve this, Boreal combines SCODA with a focusing gel containing specific oligonucleotides. The method allows perfect matches to be separated easily from mismatches, Marziali said, and either can be focused or removed depending on run parameters. The technique is capable of single-nucleotide resolution, he added.

The recently launched Aurora, available now in North America, is a spruced-up version of the previous SCODA prototype, and features a microtiter plate-sized disposable cassette that is plugged into a benchtop instrument about the size of a PC tower. The instrument is capable of 30-minute run times and can complete lysis through to sample purification in one automated step.

Marziali said that the potential disposable cost is in the range of $10; while the instrument prototypes would likely carry a price tag of about $50,000, or "hopefully cheaper."

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