Without the critical step of separation—isolating cells, nucleic acids, and proteins from biological samples—many workflows in research, diagnostics, and cell therapy would not be possible. Over the last 40 years, separation has become a common and necessary step in applications like single-cell sequencing, nucleic acid testing, CAR-T therapy, and many other areas.

Yet, despite their common use and how critical separation technologies have become, there has been far less innovation in separation as compared to nearly every other part of the biotech workflow.

The practice of cell separation has especially experienced rapid growth, roughly doubling in the last several years. This growth is attributed to the increased utilization of the immune system for the study, diagnosis, and treatment of diseases.

However, in applications like CAR-T therapy, current separation technologies contribute to the widely recognized problems of the complexity, time, and cost of the manufacturing process. In fact, advancements in immunology research and cell therapy have typically outpaced the available solutions and capability of the separation tools needed for those applications.

 Improving cell separation with microbubbles

Until recently, there have been two primary ways of separating or sorting specific cell types: fluorescence activated cell sorting (FACS) and magnetic separation, which is often referred to as magnetic activated cell sorting (MACS). These techniques are commonly used in stem cell research, oncology, and immunology. Whereas FACS uses flow cytometry instruments, MACS uses magnets and related equipment. These technologies have been the primary separation methods for decades, as the cell sorting market has had little development and improvement in their basic approach over the years.

While FACS and MACS methods are widely used, they present several challenges that limit their use. Some of these limitations include the volume that can be effectively handled, the gentleness of the method, and the number of samples that can be processed at a time.

A next-generation cell separation technology called buoyancy-activated cell sorting (BACS) that we have developed at Akadeum overcomes these limitations through the use of tiny floating particles called microbubbles, to capture and isolate target cells, like T Cells, B Cells, and even dead cells.

BACS allows researchers to perform cell separation in a simple centrifuge tube without the use of a magnet, column, or microfluidics. Microbubbles bind to targeted cells and separate from non-targeted cells by floating to the top of a sample, while non-targeted cells sink to the bottom. Researchers can then collect their desired cells from the top of the sample (positive selection) or from the bottom of the sample (negative selection).

There is an urgent and increasing need for processing large numbers of cells in large volumes with the adoption of autologous and allogeneic cell therapies. Traditional cell separation methods are restricted in volumes that can be processed, preventing researchers from efficiently sorting samples with large fluidic volumes or large numbers of cells.

However, utilizing buoyancy in cell separation, overcomes these long-standing limitations. BACS technology has no volume limitations for cell separation because it uses the uniform force of buoyancy that can be readily scaled.

Another challenge of traditional methods is that they can be time consuming and cumbersome. This limits sample throughput and risks exposing cells to unfavorable conditions, leading to long workflows and low throughput. As a result, scientists spend most of their day separating cells instead of studying them! Extended processing times needed for cell separation may also damage cells of interest in the sample. In addition, exposure to massive magnetic fields during cell separation that are well over 10,000 times more powerful than the strength of the Earth’s magnetic field, affects cell morphology and metabolism.

The growing need of high-quality cell separation has led to the emergence of new technologies that utilize microfluidics, acoustics, and filtration. However, these emerging technologies have many of the same drawbacks as traditional methods because they employ MACS and FACS.

As new developments in cell separation technology continue to arise and transform the industry, the same can be expected of scientific and diagnostic discoveries and therapeutic applications like cell and gene therapy. With researchers now able to perform cell separation more efficiently and effectively using BACS microbubbles, a new path is being forged for improvements in healthcare.

Brandon H. McNaughton, PhD, is co-founder and CEO, Akadeum Life Sciences.

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