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Home Insights & Advice Covid-19: Digital PCR company Stilla explores a novel approach for detection

Covid-19: Digital PCR company Stilla explores a novel approach for detection

by John Saunders

2020 has been quite the rollercoaster ride. Covid-19 had caught the world at unawares and quickly snowballed into a global pandemic. But the medical community has stood its ground, racing to find solutions to halt the spread of the virus.

Stilla Technologies has made a breakthrough in the field. The company’s research team has developed a Digital PCR kit combined with its system that can detect the virus.

Evolution of PCR

In the last two decades, PCR technology has made tremendous progress. A quick look at its evolutionary curve will help better understand the leap that Stilla Technologies has made with its Crystal Digital PCR.

Limit dilution PCR

In 1983 Kary Mullis discovered Polymerase Chain Reaction (PCR) and its uses to manipulate and detect nucleic acids. The 1990s saw the evolution of limit dilution PCR experiments. This process involved diluting the template DNA to such an extent that any single PCR reaction contained only a single template molecule.

Because of the multiple reactions in the limit diluting PCR, the frequency of positive and negative reactions will always follow the Poisson distribution. Based on the dilution factor, you can thus calculate the abundance of target molecules.

Digital PCR

In 1999 Vogelstein and Kinzler developed a new process and coined it Digital PCR. In this process, a sample is partitioned into numerous distinct components. PCR reactions then occur individually in each of these components. With the development in microfluidic engineering and design Digital PCR quickly balanced out its initial drawbacks of being time-consuming and expensive.

Around the mid 2000’s the researchers came up with a new Digital Array PCR method. This method utilized a chip containing 12 different panels which were partitioned into 765 6-nL chambers. This method however was limited by the increased time it took to test samples, the high costs per test and the restricted volume of reactions.

Crystal digital PCR

The real breakthrough in Digital PCR came in 2016 with Stilla Technologies’ Naica system. Naica is a unique Digital PCR platform that has high sensitivity and precision, with a 3-color target multiplexing capability. This process partitions the sample into an 2D array of microdroplets by confinement gradient.

The confinement gradient generated crystal droplets ensure homogeneity in the droplet size. The droplet size does not depend on the physical properties like surface tension or viscosity of the droplets. Crystal digital PCR™ offers an integrated workflow and multiplexing while making the whole process easy-to-use. This technology is used for nucleic acid quantification in assays, including rare event detection and infectious diseases like Covid-19.

Shortcomings of RT-PCR as a testing method for Covid-19

The central strategy to combat the pandemic is to restrict its spread through testing and social distancing. The medical workers are predominantly diagnosing patients using virological tests. These tests are based on a technology called real-time reverse transcription-polymerase chain reaction (RT-PCR).

However, due to the rapid spread of the virus, the RT-PCR technique is reaching its limit. Simply put, it just cannot keep up with the massive volumes of testing that the situation demands. The cost of this testing process was not scalable to the world population. This technique also fails to consistently detect low viral loads in cases of patients who are mildly infected.

Group testing

The world was sorely underprepared to handle such an infectious viral disease. As a result, medical institutions worldwide saw a severe shortage of reagents essential for RT-PCR. A temporary solution was to pool together individual samples for group testing. The test positivity rate for COVID-19 testing is usually below 10%. Such a low positivity rate makes it an optimal configuration for group tests.

In a group test, the samples of 8, 16, or 32 individuals can be pooled together. This pooled sample is then tested, and if the test is negative, it implies that none of the individuals in the group have contracted the disease. A positive test result would mean that one or more individuals are carrying SARS-CoV-2. The individuals from the group are then tested separately to pinpoint the carriers.

Group testing effectively reduced the amounts of reagents needed by a factor of five. However, this had its own back draws. Testing any group larger than 4 would result in a lowering of the sensitivity of the tests. This was evident from the alarming number of false-negative results.

Stilla Technologies’ novel solution

Stilla Technologies has developed an elegant solution to the shortcomings of the RT-PCR testing method by introducing a Digital PCR technology called Crystal Digital PCR. A very recent study showed that the Naica System can accurately, quickly, and reliably identify the SARS-CoV-2 virus.

Naica system is a testing approach that combines Digital PCR with group sampling. This effectively reduces the reagents required by 80% while greatly increasing the test sensitivity. The Naica system also requires fewer samples, which reduces the risks of cross-contamination.

The DIgital PCR testing process by Stilla is not only more cost-effective, but it also increases the capacity of testing by 10 times. These are significant advantages that are helping to turn the tides against the viral pandemic.

Effective application

A close collaboration with Apexbio, (Stilla Technologies partner in China) allowed the company to quickly develop a testing solution. Using its Naica System the medical professionals could promptly detect Covid-19 through digital PCR.

Since the pandemic started, more than 15 Chinese institutions (the First Affiliated Hospital of Zhengzhou University, the Center for Disease Control & Prevention, etc.) have been using this Digital PCR solution. They successfully carried out advanced testing in more than 1,200 samples from patients for whom RT-PCR tests were inconclusive.

This early deployment of the technology in combating the spread of the SARS-CoV-2 virus gave Stilla Technology a headstart and a better understanding of the range of Digital PCR’s applications. Since the end of March 2020, Stilla Technologies has also started testing its solution in several French hospitals.

Extensive research and international collaborations confirm that Digital PCR allows a more precise and sensitive identification and quantification of the SARS-CoV-2 virus. This innovative solution by Stilla Technologies (stillatechnologies.com) can scale up to meet the global expectations as the fight against the pandemic continues.

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