Koen Martens from the Institute for Microbiology and Biotechnology at the University of Bonn working at the custom-built super-resolution fluorescence microscope that he uses for his investigations. Credit: Volker Lannert
Researchers at the University of Bonn have developed a novel high-throughput method to observe and track molecules up to five times faster than current methods. The speed increase will allow future researchers to gain insights into unknown cellular functions.
Despite cellular imaging techniques existing previously, they lacked the power to accurately track individual molecules making many cellular processes remain a mystery. "Single particle tracking involves marking the molecule with fluorescent light, making it into a kind of light bulb," said Koen Martens from the Institute for Microbiology and Biotechnology at the University of Bonn.
"We then take hundreds of photos a second using a high-resolution microscope. Our 'light bulb' lights up the molecule in the darkness of the cell, allowing us to observe it and track its movement over time. This enables us to measure its diffusion and how it interacts with other cellular components."
While these methods have provided unprecedented insight into many functions within a cell, they have one glaring drawback. "It's hard to track multiple molecules at the same time," said Martens. "When their paths cross or they're too close together, you get two light bulbs merging, in effect. Then it's impossible to identify their movements."
To solve this problem, Martens created TARDIS (temporal analysis of relative distances) which runs an all-to-all analysis of the distances between two locations such as the location of the molecule in individual photographs. Published in Nature Methods, TARDIS looks at the entire sequence of movements within a cell instead of focusing on individual points such as in previous imaging methods.
By examining the entire sequence of movements, TARDIS can scrutinize all of the molecules in a cell simultaneously. "TARDIS makes the measurement process at least five times faster without any loss of information," added Martens. In the future, the researchers intend to use TARDIS to investigate the processes involved in DNA repair. "I'm especially interested in investigating how easy or difficult certain kinds of damage are to repair and how badly the DNA is damaged by a specific dose of UV radiation or chemicals," said Martens.