Technology

Technology

Cellular Highways instruments are powered by a new type of microfluidic cell sorter, VACS: vortex actuated cell sorting (patent pending).

Like in conventional cell sorting, a stream of fluorescently labelled cells is measured optically, and sorting decisions are made in real time. In our technology, cells are deflected individually by a new type of sorting mechanism: a microscopic inertial vortex in the flowing medium that is created by a thermal vapour bubble. Our technology requires no side channels, no sheath fluid and no aerosols, only one input and two outputs (sorted cells and waste) on a microfluidic chip.

By multiplexing this technology, we are able to deliver far higher sort rates than existing platforms, and thus enable new diagnostic and therapeutic applications.

  • Large Instrument

    In biosafety cabinet, load syringe with cell suspension.

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    Insert syringe in sterile Highway 1 cartridge.

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    Load cartridge in instrument, set gates.

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    Walk away and wait for sort to complete.

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    Remove cartridge, take to biosafety cabinet, remove output tubes.

Cell sorting with an inertial vortex

VACS uses a thermal vapour bubble actuator, akin to the actuator developed in thermal inkjet printing, to deflect cells. These actuators have some attractive properties for microfluidic applications. The actuator is a very small (~100 µm) electrical microresistor – much smaller than ultrasonic actuators, for example – and can be made by standard MEMS manufacturing processes on a glass or silicon substrate. It is driven directly by an electrical pulse without the need for external transducers, such as external electromagnets or piezo-actuators.

The displacement produced by a single thermal inkjet-style actuator is extremely small and short-lived, however. Using thermal inkjet-style actuators to deflect cells is therefore a significant challenge. To deflect a cell, it is necessary to create a displacement greater than the diameter of the cell. Moreover, the time scale of the displacement needs to match the transit time of the cell through the junction between the input and output channels. If the displacement is too short-lived, cells are pushed one way and then the other before they enter the sorting channel, and are therefore not sorted.

Previous researchers have solved this problem by: (1) placing the actuator in a side channel to focus the displacement in a smaller area, (2) using many thermal inkjet-style actuators in parallel to increase the displacement amplitude and duration, or (3) employing a high-power laser rather than an electrical microheater to create the bubble. However, these approaches add complexity to the sorter chip and make it difficult to envisage a practical, low-cost or parallelisable cell sorting instrument.

We decided to position a single thermal inkjet-style actuator at the side of the main sorting channel, thus avoiding the need for additional side channels. At the same time, the actuator is far enough from passing cells so that the cells don’t feel the heat from the vapour bubble – the cell viability is unaffected.

With the actuator positioned at the side of the main channel, we then required a way of converting the small, short-lived displacement generated by the vapour bubble into a large, lasting cell displacement.

We achieved this by designing a channel geometry that creates a transient vortex from the expansion and collapse of the bubble. In this configuration, the thermal vapour bubble creates an inertial vortex at a well-defined position in the stream that flows downstream with the cell to be sorted, thus causing a permanent cell displacement.

Thanks to accurate simulations of the fluid dynamics of this system, we were able to design and test the inertial vortex in silico. The animations below show how an inertial vortex deflects individual cells.

We then built a complete fluorescence-activated cell sorting system, implemented in a demonstrator rig, to show that inertial vortex sorting works. The sorting process is too fast for even high-speed imaging, but a strobe light allowed us to visualise and verify sorting events.

Simulation of the inertial vortex sorter in 2D

Deflection of a cell in a stream by an inertial vortex, showing the total flow (steady stream left to right, from input to outputs).

Inertial vortex flow visualisation

The same simulation, showing the transient flow only: the creation of the inertial vortex and its deflection of a single cell can be seen clearly.

Verification of positive sort events

Experimental verification of the sorting system by double-strobe imaging. The video shows a compilation of independent double-flash strobe frames (rather than a real-time video).

Advantages of VACS

Our core technology is fast, small, suitable for multiplexing and highly manufacturable.

In the demonstrator rig, VACS achieves the following:

  • Equivalent droplet rate / switching speed of 43,000 / second
  • Maximum actuation rate of 50,000 / second
  • Sustained deflection rates currently around 4000 / second
  • Error rates for 10 µm particles: < 0.01% (false positives), < 0.5% (false negatives)

Our core technology is also extremely small. The dimensions of the device on chip is 1 mm x 0.25 mm, without the need for an external actuator or side channels. Therefore, our core technology is attractive for multiplexing on chip or integration with lab-on-chip workflows.

Finally, our basic cell sorting chips are highly manufacturable because they are made with standard MEMS processes and materials and incorporate only a single layer of microfluidics.

The sort envelope is the key measure of performance of a particle-sorting technology. This function of time or space along the stream shows the basic ability of the technology to pick particles precisely and avoid false positives or false negatives. Our envelope width is 52 μm (or 23 μs).
Our core device is both small and simple, incorporating only an input, two outputs, no side channels, and a 250-µm-wide device in the centre. The actuator and electrical connections are made by standard MEMS thin film processing techniques on a glass substrate, and a single microfluidics layer is bonded to the glass.
Scientific paper in Lab on a Chip

Scientific paper in Lab on a Chip

We published full details of our core technology in the scientific journal Lab on a Chip. The main paper shows the physics of the device, demonstrates sorting of beads and peripheral blood mononuclear cells (PBMCs), and compares the results with cell sorting theory. The supplementary information details our laboratory demonstrator rig, which incorporated a single laser (488 nm), lenses, filters, detectors and fast electronics to measure fluorescence, forward scattering and backscattering to decide which cells to sort in real time. Evaluation of the technology was done in collaboration with the GSK Clinical Unit Cambridge

“Cell sorting actuated by a microfluidic inertial vortex”, R.H. Pritchard et al. Lab Chip, 2019, 19, 2456-2465

16X Parallel Sorting

16X Parallel Sorting

We built a functional demonstrator of a 16X parallel version of the technology. The 16X system sorts living cells much faster than any device has ever achieved in the past. The instrument is based on the same principles, but consists of 16 VACS devices on a 4 x 4 grid of pitch 1 mm. We showed a video of 16X parallel sorting at our talk at CYTO 2019, Vancouver.

  • Sort envelope rate: 2.5 billion / hour
  • Practical cell processing rate: 0.7 billion / hour

The fastest cell sorter in the world: 16x parallel VACS from Cellular Highways on Vimeo.