John He, Mechanical Engineer at Cellular Highways

During the design of the Highway1 cartridge we encountered many engineering challenges, due to the novelty of our core technology, the cytometric measurement requirements, and the aims of cell therapy.

From the outset, we know that the cartridge needed to be sterile, fully enclosed and low cost. At the same time, it had to be easy to use and integrate conveniently into both research and good manufacturing practice (GMP) workflows. Since it has to be single-use and closed, there could be no clogged nozzles, no cleaning, and no carry-over or contamination. In addition, the input vessel had to be large enough to process typical cell populations in a single run without the need to use multiple cartridges.

To meet these challenges, we did much research and consulted a wide range of stakeholders – alpha testers, cell therapy manufacturers, regulators, users and other stakeholders – before we settled on a definitive user requirement specification.

Naturally, users face many decisions during sample preparation, including choice of media and reagents as well as cell handling steps such as thawing, washing, pre-enrichment and resuspending. For some users, most of these are already possible as closed processes. But for many, these steps are still open. So for the first release of the cartridge we assumed that sample preparation would still be carried out using aseptic techniques, inside a biosafety cabinet, isolator or cleanroom. The input syringe is designed to fit conveniently into these workflows. And to enable easy sample loading, the syringe is supplied with a pre-attached filler tube and in-line cell strainer.

We decided to initially develop two cartridge versions: one for research use only (RUO) and one for GMP. For the RUO version, we chose to employ centrifuge tubes as the output vessels, to ensure easy washing or transfer of cells to downstream processes. To ensure functional closure a sterile vent filter was selected that achieves over 99.99978% Bacterial Filtration Efficiency (ASTM F2101). For the GMP version, the output vessels will be bioprocessing bags, so that the user could use aseptic tube welding to achieve fully closed downstream processes.

We still have some more work to complete before Highway1 can be launched. A good example of this is the cartridge frame, with which the user handles and mounts the cartridge in the instrument (the cartridge frame does not contact the sample). For beta users, the frame is 3D printed rather than injection moulded to allow us to do more human factors testing with beta users: 3D printing allows us to rapidly iterate the finer details of the frame based on user feedback. Once we are confident the cartridge is as user friendly as possible, we will finalise the frame design and begin injection moulding.

We are also in the process of setting up cleanroom mass production of the cartridges. Here our goal is to ensure the cartridges are manufactured and tested to meet the relevant regulatory and safety requirements before we place them into the hands of our users.

After the initial release of Highway1, we plan to release a further cartridge variants to support cell sorting for specific applications. Our priority are cartridges for depletion sorting (negative selection), followed by cartridges for sorting different cell sizes, including smaller cells such as microbes, and larger cells such as differentiated iPSCs, macrophages and plant cells.

Beyond Highway1, there is a lot of exciting work in store for Highways16 – parallelising the technology to achieve 16 times the throughput! Although the core technology has been proven (https://www.mdpi.com/2072-666X/12/4/389), much engineering and development still lies ahead –  not least because Highways16 cartridges will have to handle much greater volumes. The input and output vessels therefore require scaling up to match the increased throughput. We also intend to focus the cartridge design towards GMP / therapeutic applications and making further workflow improvements, such as using bioprocessing bags for the input instead of a syringe. This will enable the entire cell sorting procedure to be performed as a closed process.

About me

Hi, I’m John. I’m a mechanical engineer at Cellular Highways, and a recent graduate from Imperial College London in 2019. Working at Cellular Highways has allowed me to learn a lot about optical instrument design, and to gain a passion for cutting edge technology in high impact applications. I joined the Cellular Highways team in June 2020; I was attracted by the opportunity to work on a highly complex instrument that integrates biology, optics, electronics and microfluidics which could revolutionise cell therapy manufacturing.