NEW YORK – Stilla Technologies, a French digital PCR developer, recently debuted a new digital PCR system called the Nio+. The single-instrument system supports a new method using color combinations in seven channels to multiplex up to 21 targets and can process up to 48 of the firm's 16-well Ruby chips per day.
The Nio+ has been in development for about three years with the goal of taking a "leap forward" in digital PCR usability, according to Stilla CEO Rémi Dangla. While the single benchtop instrument incorporates a large touchscreen interface and internal design features enabling continuous feeding of experiment plates, it also features "the French touch" in its aesthetics, Dangla said in an interview.
Stilla officially launched the instrument last week with a 30-minute video presentation demonstrating features of the system. The video also included comments from early-access users like Wim Trypsteen, a cofounder of the Ghent University Digital PCR Consortium in Belgium who frequently beta-tests digital PCR systems as part of his work.
Trypsteen noted in a follow-up interview that there are only a handful of single-instrument digital PCR systems currently available commercially but said that, in his opinion, a single-instrument system and additional features focused on ease of use are becoming essential.
In the digital PCR space, a single-instrument design was initially pioneered by the Qiagen QiAcuity and Absolute Q from Thermo Fisher Scientific. After the QiAcuity was launched, "people were like, wow, this is where we should be — this is the standard now and people will want to have ease of use in the future," Trypsteen said. "It's a must-have."
Dangla also said that this ease of use is important across all markets, from wastewater testing to biopharma development.
"Customers shouldn't have to invest time in becoming a technology expert," he said. "They need to spend that time on analyzing samples [and] developing analytical solutions for their customers, and so we want to offer those customers something that you just plug it in your lab, have one day of training, and then you're pretty much autonomous to use the machine."
For Stilla, the approach to improving usability began at the first interaction that users have with the instrument, Dangla said.
Accessing the software to start operating most digital PCR systems involves a mouse and a keyboard, he said, but he asked his design team to think about how end users actually approach an instrument.
"The person in the lab will walk to the machine with the [dPCR experiment] plate in his or her hand," he said.
So, for the Nio+, inserting a plate starts the workflow. "Without touching the instrument any other way, it will automatically recognize your plates, scan the QR code, and then there's a pop-up asking you: 'What experiment do you want to run on those chips?'" Dangla said. The user then selects a preprogrammed experiment, hits "Go," and walks away.
In addition to obviating the need to move plates between droplet-generating and thermal-cycling instruments, the Nio+ also allows for random access and so-called continuous loading of experiments.
The system accepts plates with up to three of the company's Ruby chips. Each Ruby can accept 16 samples, meaning a plate of three can contain up to 48 samples. Up to eight of these chip plates can be loaded in the system for processing, and as plates finish their run, additional plates can be added. According to the launch video, the system can process up to 768 samples per day using this continuous loading approach.
Within the Nio+ are multiple robotics systems that transport the plates in an asynchronous manner from the loading area to the thermal cycler, Dangla said.
There are two thermal cyclers in the instrument, but "each thermal cycler unit has three subregions that are independently controlled, so each subregion can have its own PCR program," Dangla noted.
In addition to running many different experiments simultaneously, the Nio+ has features that enable higher multiplexing in each well.
Stilla's most recently launched system, the Naica Prism 6, has six optical channels, but Nio+ adds a seventh that detects large Stokes-shift dyes. Other digital PCR instruments on the market to date have six or fewer optical channels.
While users can employ all seven of the Nio+'s optics channels to detect seven fluorophores per well, the firm has also developed a multiplexing method called "color combination" for higher-order multiplexing. This approach also benefits nonlinearly from the addition of the seventh channel.
With this method, each target is encoded by the presence of two fluorophores per droplet. For example, the first target could be linked to a blue and a green dye, the second could be blue and red, the third blue and yellow, and so on. "If you have six channels and you do color combination, you have 15 combinations of colors and can actually detect 15 targets," Dangla said, but, "if you have seven channels, then you can go up to 21."
Droplets that have more than two colors must be discarded from the analysis using this approach, as they are ambiguous.
"But, for the vast majority of applications that we have in mind, especially liquid biopsy applications, the concentration of the targets is going to be low because you're looking at rare event detection, so the probability of having many triple-positive [or] quadruple-positive droplets in your assay is very, very low," Dangla said.
Trypsteen also emphasized the ease of use of this color-combination approach, noting that it is more straightforward from an assay optimization point of view, as compared to error ratio or amplitude-based multiplexing. Detecting 20 targets with six or fewer color channels using other methods would actually be "way more challenging than what is possible now with seven colors," he said.
The large Stokes-shift dye seventh channel can also be used for reference gene detection, leaving 15 options for a panel of rare targets, Dangla said.
Cost is also a critical element in most lab testing these days, and Dangla said many customers wishing to take on digital PCR are coming from the low-cost qPCR space. The firm now plans to offer its legacy Naica system at a lower cost for those venturing into dPCR, while offering the Nio+ at a "competitive" pricing compared to other, similar systems, Dangla said.
Overall, for the Nio+, Dangla said he has pushed the Stilla R&D teams to think hard about ease of use and also consulted intensively with end users.
"We're developing powerful technology, but it makes no sense if the technology is powerful but can only be used by a handful of experts in the world," Dangla said. "We really thought about making powerful technology that is also very easy to use."