In cancer research, it all comes down to a single cell.
Over the past decade, cancer researchers have taken to the fact that a single cell from a tumor can be used to perform molecular analyzes that reveal important clues about how cancer developed, how it spreads and how it can be targeted.
With this in mind, a team of Brown University researchers developed an advanced way to isolate single cells from complex tissues. In a study published in Scientific reportsthey show how the approach not only results in high-quality, intact single cells, but also outperforms standard isolation methods in terms of labor, cost, and efficiency.
The challenge was to develop technology that would allow researchers to more quickly and easily isolate cells from biopsied cancer tissue to prepare it for analysis, said Anubhav Tripathi, study author and director of biomedical engineering at Brown.
“Technologically, there’s nothing like it on the market right now,” Tripathi said. “This technology will be useful for those looking for answers using genomics, proteomics, transcriptomics – it will not only make these diagnostic and therapeutic studies easier, but also save researchers time and effort.”
Tripathi added that beyond clinical applications, the technology will be useful in biomedical applications such as tissue engineering and cell cultures.
Single-cell analysis uses advanced sequencing techniques to obtain genetic profiles of individual cells. This is particularly applicable to cancer tissues, where rare mutations can drive metastasis and treatment outcomes. A major limitation to the clinical translation of single-cell analysis is the difficulty of isolating single cells from complex tissues, said co-author Nikos Tapinos, associate professor of neurosurgery and neurology at Brown.
Tapinos describes a typical workflow using the example of a brain tumor: A portion of the tumor will be removed in an operating room and taken to a laboratory. There, researchers would use a process involving enzymes to extract nucleic acids from massive tissue samples and then perform mass genetic sequencing.
This process results in low resolution, potentially inaccurate genetic readings and poor detection of rare cell types, Tapinos said. The consequences of losing this information can be profound, he noted, including the possibility of misdiagnosing a patient, creating a significant delay between the time the tumor is removed from the patient and the cells are ready for RNA sequencing.
“There is a huge need for technology that allows tissue to be removed from the patient within minutes, results in viable, healthy single cells from which RNA can be isolated,” Tapinos said. “That’s exactly what this new process does.”
The advantages of electricity over enzymes
In the new process, a tissue biopsy is placed in a fluid-filled vessel between two parallel plate electrodes. Instead of enzymes, electric field fluctuations are applied to create opposing forces in the liquid. These forces cause tissue cells to move in one direction and then in the opposite direction, causing them to separate cleanly or separate from each other.
This approach was invented by study author Sel Welch, a fourth-year Brown Ph.D. candidate in biomedical engineering in Tripathi’s lab.
“Dr. Tripathi has done a lot of work in his lab using electric fields and microfluidics,” Welch said. “After seeing how electric fields could be used in other diagnostic applications, we had the idea to do something unique with the electric field that had never been done before. Based on the existing body of research on the manipulation of biological particles, we formulated a hypothesis about how this would work.”
The new process resulted in dissociation of biopsy tissue in just 5 minutes—three times faster than leading enzymatic and mechanical techniques described by Tripathi and Welch in a previous study.
The approach also resulted in “good tissue dissociation into single cells while preserving cell viability, morphology and cell cycle progression, suggesting utility for tissue sample preparation for direct single cell analysis,” the study concluded.
According to the researchers, the new approach is at least 300% more efficient than even the most optimized techniques using simultaneous chemical and mechanical dissociation.
Another advantage of the process, Welch said, is the compactness of the device they created: “In a traditional workflow, you have to use several different lab tools, such as a centrifuge tool, each of which costs several thousand dollars. single-cell sample preparation requires only one device.”
The research team has applied for a US and global patent for the device and its associated methodology, with the assistance of Brown Technology Innovations, the university’s technology transfer office.
The samples used in the study were bovine liver tissue, triple-negative breast cancer cells, and human clinical glioblastoma tissue. The research team is now refining the technology and developing a device that will be able to quickly and efficiently process many different types of tissue biopsy samples on a small scale at once, at a very low cost.
The new study explains the scientific basis for the process, Welch said. “We are now working on a new device that is specifically geared towards creating this highly optimized system to exploit this physical phenomenon.”
“A researcher will be able to simply push a button and within minutes have the individual cells they need for analysis,” Tapinos said. “It’s really amazing.”
Harry Yu, a graduate student at Brown University, and Gilda Barabino, president of the Franklin W. Olin College of Engineering, also contributed to this research. The work was supported by funding from PerkinElmer, where Tripathi serves as a paid scientific advisor/consultant and lecturer.