In 1885, German biologist Wilhelm Roux took part of the brain from a chicken embryo and placed it in warm saline solution, keeping it alive for several days. Simple as the experiment was, it seeded the idea that biological tissue could be maintained, or cultured, outside the body, given the right conditions.
Over the years, scientists have refined their methods for keeping various tissues—and even individual cells—alive in petri dishes or cell culture plates. The ability to see, stain, probe and genetically engineer living cells in vitro has paved the way for a deeper understanding of how biological systems tick and formed the basis of screening tests for drugs against a plethora of diseases.
However, in vitro cell culture and the accompanying downstream analyses remain manual and tedious due to limited choices in methods for washing cells. Reagents are repeatedly added to cells and rinsed away with manual processes, sometimes resulting in the loss of biological material. At the same time, assay outcomes are highly dependent on the experience and technical capability of the researcher performing the experiment.
“There is currently a significant void in ensuring the consistency of cell analysis because practically all cell analysis is performed manually, leading to variation between operators, locations, times and so on,” said Namyong Kim, CEO of bioinstrumentation company Curiox Biosystems. “I believe that these problems can be solved with automation and quantification.”
Kim added that cell analysis is becoming more important in emerging areas of research, including the profiling of small populations of cells such as tumor-infiltrating leukocytes in dissociated tumor samples, or the sequencing of single-cell genomes. In these experiments, a robust and accurate system for handling and processing cells is critical, he said.
Bringing down walls
Kim joined A*STAR’s Institute of Bioengineering and Nanotechnology (IBN) in 2004 with ideas for creating an automation system for cell analysis. Before that, he had been working with early-stage high-tech ventures and multinational companies in the US. His industry experience in developing biological assay platforms led him to realize that the assay format had a strong influence over the way downstream processing and analysis could be carried out.
Many high-throughput biological assays involve placing cells in plates consisting of 96 wells separated by walls. When reagents are added or removed from these wells, convective currents and turbulence arise, stressing cells and causing some of them to be washed away during rinsing steps.
Hence, his team at IBN conceived the idea of a wall-less plate that could still accommodate 96 distinct samples. “We developed a super hydrophobic (water-repellent) surface containing hydrophilic (water-‘loving’) spots positioned at regular intervals. Each hydrophilic spot stabilizes an aqueous droplet, giving rise to an array of sample droplets on a plate. This allows us to wash the samples by unidirectional laminar flow without convective effects or turbulence,” Kim explained.
Harvesting the cells from the wall-less plate was also easier and less damaging to the samples. Kim noted that the use of laminar flow in lieu of centrifugal collection led to better consistency and cell viability, which in turn allowed for better resolution of cell populations and more effective debris removal—crucial parameters affecting data quality. The team eventually settled on two variants of wall-less plates: the DropArray and the Laminar Wash plate. These would go on to become the core technologies of Curiox Biosystems when Kim spun off the company from A*STAR in 2008.
Machines make light work
An improved assay format was only half of the equation. The next hurdle was to create a platform for dispensing reagents and performing the washing steps of assays automatically.
Working with engineers, Kim’s team devised precise computer-controlled systems consisting of fluidics and robotics to allow researchers to place their DropArray or Laminar Wash plates into a machine, load the appropriate reagents, input the desired protocol, then just walk away and let the machine handle the rest.
Curiox Biosystems has since launched a series of instruments and associated products enabling the laminar flow method in handling cells, including its Laminar Wash AUTO1000 workstation, a walk-away, fully automated cell analysis system.
Curiously, Kim commented that developing an impressive range of technology was not the most difficult part of his journey from scientist to CEO—the biggest challenge was the application of the technology to meaningful biological assays. “It was not easy to find a counterpart in the field of biological sciences that was as excited about our technology as we were,” he quipped.
Leveraging A*STAR’s contacts, Kim managed to secure collaborations with the Singapore Immunology Network (SIgN) to validate the Laminar Wash technology for cell analysis and DropArray technology for protein analysis. For example, SIgN implemented Curiox Biosystems’ DropArray system on the Institute’s clinical immuno-monitoring platform. The researchers showed a five-fold reduction in assay reagent volumes and clinical sample usage, and 65 percent savings in overall clinical assay costs.
Another partnership with the Institute of Molecular and Cell Biology (IMCB) involved the optimization of the DropArray system for RNA transfection of cells in suspension. The findings not only helped refine Curiox Biosystems’ technology, but also facilitated the discovery of fundamental biological mechanisms at IMCB.
Innovation crossing borders
“Today, our DropArray and Laminar Wash technologies are being used by more than 80 percent of the 20 biggest biopharma companies, with many owning multiple units of our products,” Kim said. “Our customer base ranges from R&D and process development to cell manufacturing and quality control, in both industry and academia.”
Clinical labs have also adopted Curiox Biosystem’s platforms because of the benefits of better and more consistent data, which is of crucial importance in organ transplant donor-recipient matching, he added.
The company’s sustained success has caught the eye of investors. In 2010, Curiox received a large corporate investment by the Zicom Group, a Singaporean-Australian conglomerate listed on the Australian Stock Exchange. More recently, in October 2019, Curiox Biosystems raised US$15 million in series B funding led by top-tier bio venture capitals in South Korea. The latest injection of funds will be used to scale up and accelerate the commercialization activities of Curiox Biosystems globally.
Reflecting on the milestones that Curiox Biosystems’ has achieved over the past twelve years, Kim acknowledges A*STAR’s role in helping the company find its footing in a competitive global landscape of assay platforms. Even as it expands abroad, it continues to engage with A*STAR’s research institutes.
“We are still in active collaboration with A*STAR—through our ongoing research collaboration agreement with SIgN, we aim to harness the strengths of Laminar Wash technology to reduce cell loss, raise the viability and yield of cells, as well as increase the resolution of distinguishing immune subpopulations compared to conventional staining protocols,” he said, adding that the findings from such studies will help to improve diagnostic immune-profiling, with implications for immunology and immunotherapies for cancer.
“Our work with SIgN has also been awarded the prestigious GAP Fund, which is governed by A*STAR's A*ccelerate. This is another testimony to the strong support available to local emerging companies,” Kim concluded.