There is a particular kind of loss that every molecular biologist has to make peace with. Reading a living cell’s RNA profile usually means it gets destroyed during the extraction process. The moment a sample is lysed to access its inner molecules, that population is gone, along with any chance of asking it a second question.
“Important questions about how the same cells change over hours or days, like tumours adapting to a drug, have to be tackled indirectly,” said Giulia Adriani, Principal Scientist at the A*STAR Skin Research Labs (A*STAR SRL). “Researchers typically need to test different cell samples at different time points and assume they were behaving in a comparable way.”
Adriani, in collaboration with Andy Tay, Presidential Young Professor at the National University of Singapore, set out to develop a gentler RNA sampling platform that could keep the cells alive. The method, called nano-electroextraction (NEE), uses millions of hollow nanostraws and applies brief electrical pulses to the cell membrane to open tiny holes that then quickly reseal, much like “sipping” the RNA molecules from the cells.
A major challenge was extracting enough RNA without stressing the cells or altering their gene activity. Ying Jie Quek, a former A*STAR Graduate Scholar, fine-tuned the strength and duration of the electrical pulses, finding that the optimal settings varied for different cell types. “This way, the same community of cells can be revisited repeatedly, without needing a fresh sample at every time point,” said Adriani.
The researchers validated NEE on primary immune cells, primary lung fibroblasts and a lung cancer cell line, benchmarking results against conventional RNA extraction techniques. Over three days, the platform accurately tracked cellular changes, such as their developmental progression or adaptive reprogramming of biological functions, by capturing thousands of genes per replicate while keeping cell viability above 80 percent.
NEE also held up in more complex three-dimensional (3D) cancer spheroids, with their structure left intact and gene activity largely undisturbed. Despite mainly sampling from the outer spheroid layers, the approach picked up meaningful signalling pathway and proliferation-linked changes that matched results from destructive lysis methods.
“This suggests that NEE is robust enough to work not only with simple, flat cell layers, but also with more tissue-like systems, opening the door to its potential use in organoids and other 3D models,” said Adriani.
Looking ahead, the team hopes to push NEE deeper into 3D tissues to unlock insights into how micro-environmental differences can contribute to tumour progression and treatment response. They also aim to extend their approach to organoids or tissue slices to track both temporal and spatial dynamics of RNA expression in more complex, human-relevant models.
The A*STAR-affiliated researchers contributing to this research are from the A*STAR Singapore Immunology Network (A*STAR SIgN).
