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Biodesix Turns to Digital PCR to Maintain Round-The-Clock COVID Testing

NEW YORK – How can a commercial diagnostic lab maintain a 24-hour turnaround time for COVID-19 testing? One way might be to scale up test platforms and technologies that don't use the same supply chain as other major diagnostics systems. Biodesix has done just that, using Droplet Digital PCR from Bio-Rad Laboratories to support returning thousands of test results each day in Colorado.

Unlike RT-qPCR, digital PCR does not require calibration to a standard curve and is more tolerant of PCR inhibiting substances. Nucleic acids in a sample are split into tens of thousands of individual ddPCR reactions, each ideally containing a single copy of the target, and the quantitative readout is essentially binary – a fluorescent glow, or not, in each tiny drop, that is detected with a fluorimeter.

Biodesix formed a partnership with Bio-Rad in March for the SARS-CoV-2 test, and since then, "They've done a phenomenal job responding to the crisis," Carolyn Reifsnyder, director of global product marketing at Bio-Rad's Digital Biology Group, commented in an interview.

The Bio-Rad ddPCR technology had been shown earlier this year to be potentially useful in test-of-cure assessment. In studies using patient samples in China, the test had reduced false negative results in samples with low viral load.

Biodesix launched its testing on April 8 and is continuing to maintain a 24-hour turnaround time for diagnostic testing, processing approximately 3,000 tests each day, according to Gary Pestano, chief development officer at the Boulder, Colorado-based company. The firm uses the Bio-Rad QX200 and QX One instruments for its testing services.

Bio-Rad received Emergency Use Authorization from the US Food and Drug Administration for its ddPCR SARS-CoV-2 test in early May.

Its flagship digital PCR instrument is the QX100, which was introduced in 2012 and developed from technology acquired when Bio-Rad acquired QuantaLife. The QX200 followed in 2013, and the firm launched an automated droplet generator the following year. The QXDx instrument was cleared by the US Food and Drug Administration in February last year and is a designated instrument for lab testing with a planned menu, in the pre-COVID era at least, of oncology and prenatal testing assays.

Most recently, Bio-Rad debuted the QX One late last year. That system puts the droplet generating mechanism, amplification, and droplet reader, in the same instrument and automates the workflow. It has a five-plate capacity and can analyze up to 480 samples a day, detecting four colors simultaneously.

On a call to recap the firm's second quarter earnings, Bio-Rad Life Science Group President Annette Tumolo, said that previously the firm had not focused its ddPCR technology on infectious disease.

"I think what has emerged is how it can be quite useful in this area, in virology in particular," Tumolo said, adding that there may be a lasting impact going forward.

Indeed, Reifsnyder said that Bio-Rad is always evaluating new market opportunities where ddPCR offers a differentiated and compelling solution to users. For example, in addition to the COVID-19 infectious disease test, Bio-Rad has also been expanding on collaborations in the US and internationally to bring the advantages of digital testing to wastewater monitoring.

Biodesix is a specialty reference lab that has traditionally focused on lung disease testing, specifically liquid biopsy for lung cancer, Pestano said. The pivot from oncology to infectious disease may not be as extreme as it might appear on the surface, however, because the firm has extensive relationships with pulmonologists, who have themselves been called up to treat COVID patients.

Biodesix adopted the Bio-Rad ddPCR systems for its rare variant testing of circulating tumor DNA in early 2015. "We've had a five-year history with Bio-Rad and Droplet Digital PCR with liquid biopsy," Pestano explained.

And, the fact that the company already had the ddPCR systems "turned out to be quite favorable," Pestano said, and "is the reason are able to keep doing what we're doing in such a rapid time."

Biodesix has two CLIA labs in the US. The one in Boulder offers ddPCR-based COVID-19 testing, while the one in De Soto, Kansas is offering antibody testing for the coronavirus.

When the COVID-19 pandemic initially hit the US , the firm was undecided about whether to shift gears from its core business, Pestano said. "I think the feeling back then was that the large labs would handle the testing," he said.

But as the pandemic developed, Biodesix realized, "We have capacity, we have instruments, we have knowledge, we have the people, and we have a partnership with Bio-Rad," so the firm started with a strategy to focus on Colorado, and "join in the fight," Pestano said.

Early on, Bio-Rad and Biodesix both pursued EUA submission. Bio-Rad's platform was approved along with an assay, and FDA gave the feedback that since the Biodesix assay was so similar, the firm should just fall under the Bio-Rad EUA, Pestano said.

In March and April, "as New York was really hot and parts of California, as well," in terms of a coronavirus spike, Pestano said most other diagnostics instruments and reagents were in allocations and preferentially going to hotspots and frontline worker testing. The national supply of reagents and plastic consumables has also been constrained for some time, without clarity about when that may ease.

Pestano estimated that 90 percent of labs are using RT-qPCR, and "if you are in any of the instruments with the highest demand, and you're not prioritized, you're going to feel some pain," he said.

"The commercial lab was really not prioritized in the list of who was going to get instruments and reagents," Pestano said.

"That became fortunate for us because those reagents are still in short supply, and that, in part, is leading to delays. Because we use the Bio-Rad platform, we are in a good position because we are not competing for reagents that are now allocated to major labs and hotspots."

Biodesix tried to go after purchasing other instruments and also develop a RT-qPCR workflow in parallel. It purchased a Thermo Fisher Scientific KingFisher platform for sample extraction, but the instrument didn't arrive until three months later. "We have already gone to market and have our lab processes in place, so we have to figure out when we have time to put that instrument into the workflow."

The Bio-Rad test uses the US Centers for Disease Control and Prevention primers and probes, and was initially authorized for use on the QX200 and QXDx with samples extracted using the ThermoFisher MagMAX Viral/Pathogen Nucleic Acid Isolation Kit or the QIAamp Viral RNA Mini Kit, according to the instructions for use.

Biodesix has adapted the workflow to incorporate a 96-well, column-based extraction method from Zymo Research, Pestano said.

"From a workflow perspective, having everything in a 96-well format just made everything easier," he said. And the Bio-Rad workflow does the reverse-transcription and PCR in the same tube, so it is a one-step reaction, he added. "That is a real time saver. My team loves it."

Besides supply chain management, another somewhat unexpected requirement for fast testing turnaround has turned out to be the accessioning process.

Pestano said that Biodesix learned accepting larger orders with repeat needs is easier to manage, since smaller accounts often send samples with handwritten manifests and tubes labeled with sharpies.

"The logistics of manifest reconciliation is one of the biggest bottlenecks in turnaround time," he said. The firm now trains customers on how to create a clean manifest before the samples arrive.

Biodesix continues to focus on its core business, too, and the firm recently purchased new Bio-Rad instruments, as well.  It has a maximum capacity to test 4,000 samples, and with optimization that could be boosted to 5,000 per day, Pestano said, particularly since the lab is now transitioning to 24/7 operations.

The firm uses the Bio-Rad QX200 and the QX One, with the latter now being applied to night shift testing because samples can run in an automated fashion overnight, "without the need for a graveyard shift," Pestano said.

Biodesix offers a national testing service program called WorkSafe that supports employee testing, and through this program, it provided all the testing to enable the continuation of the 2020 Major League Lacrosse season.

Locally, Biodesix is working with large hospital networks in the state, including Boulder Community Hospital and others, Pestano said, particularly for patients requiring quick results prior to elective surgeries. Last week it announced it will provide testing for the faculty and staff returning to Colorado State University's campus for the fall term beginning August 24.

The firm is also partnering with the State of Colorado to provide residents there with testing. In a news conference late last month, Governor Jared Polis expressed his ire that, due to the national testing backlog, it had been taking 10 to 12 days for people in Colorado to get COVID-19 lab results. "That is unacceptable; it is not helpful at all," he said.

So, "We in Colorado are going to be taking matters into our own hands," Polis said, in part by bringing additional in-state private lab partners online.

Colorado has ramped from 2,000 tests per day in mid-April to 10,000 per day in late July. The state is partnering with local hospitals and public health, and has contracted with in-state private labs. Now, Biodesix will supply 700 test per day, Polis said in the news conference, while a lab called UniPath will test 500 samples each day.

The state has also made agreements with out-of-state private labs, including Mako Medical, Curative, and ACTG Network for a total of 6,200 additional tests per day, and has an ongoing relationship with South Korean assay maker SolGent to provide additional supplies.

Using this broad approach, "We're going to get it done ourselves," Polis said.

Polis also said in the news conference that the state has contracted with Colorado State University to provide testing for testing in long-term care facilities. And, in addition, the Colorado state lab has "invoked authority from the FDA to cut through red tape and approve novel tests for use in Colorado."

The state is evaluating a saliva-based RT-LAMP screening tests from University of Colorado, Boulder, Polis said. The test was recently described in a medRxiv, and includes a sodium hydroxide pre-treatment that the researchers claim reduced false negative results that can occur when the pH-sensitive dye in the colorimetric RT-LAMP interacts with acidic saliva samples. 

Testing demand is expected to be high for some time to come. "There will come a day when we're back to normal --- and we'll all hug one another and burn our masks in a big bonfire --- but it is not today, it's not tomorrow, it's not next week, [and] it's not next month," Governor Polis said.

And in the meantime, Pestano said a community spirit that is leading to progress. "I've been in the diagnostics industry for 30 years, and I've never seen people be as helpful to each other in development of assays," he exclaimed.