Here's good news for researchers working with high-resolution fluorescence microscopy: For the first time, biocompatible molecular rulers can be used to calibrate the latest super-resolution microscopy methods. Recent advances in super-resolution microscopy have enabled optical resolution to reach a few nanometers, comparable to the size of cellular molecules. However, validating this resolution on cellular components such as multiprotein complexes has been a challenge due to the lack of biomolecular reference systems capable of precise dye labeling at nanoscale distances.
A team of scientists led by Dr. Gerti Beliu and Professor Markus Sauer from the Rudolf Virchow Center for Integrative and Translational Bioimaging at the Julius-Maximilians-University (JMU) in Bavaria, Germany, has now provided a turning point. They published new biocompatible molecular rulers, PicoRulers (protein-based imaging calibration optical rulers) in the journal Advanced Materials.
The research team successfully built these custom molecular rulers using genetic code expansion and click chemistry. They can be used as precise biomolecular reference structures in fluorescence microscopy.
PicoRulers are based on the three-part protein PCNA (proliferating cell nuclear antigen), which plays a central role in DNA replication and repair. The protein has been modified by introducing unnatural amino acids at precisely positioned positions, allowing fluorescent dyes or other molecules to click specifically onto it with minimal linkage errors.
This allows researchers to test the resolution of the latest super-resolution microscopy methods on precisely defined cellular biomolecules with unprecedented precision.
Markus Sauer said enthusiastically: "The ability to resolve real biological structures at the sub-10 nanometer level marks a new era in bioimaging technology. Compared with previously used artificial macromolecules, our PicoRulers are not only biocompatible. They also achieve unparalleled test resolution accuracy under real-world conditions."
"The scope of applications of this technology goes far beyond the boundaries of traditional microscopy," explains Gerti Beliu. "Our PicoRulers are not only a tool for more precise measurements, they also open the door to a deeper and more detailed study of the complex processes taking place within cells."
In the long term, further development of PicoRulers could transform biological and medical imaging with molecular resolution. PicoRuler enables the validation and improvement of the resolution potential of new super-resolution microscopy methods on biological samples for the first time. This makes them valuable tools for future elucidation of the molecular organization and interactions of biomolecules in cells.