Biomedical imaging is an important tool in science because it allows scientists to see what may not be visible to the human eye. Using light within the visible spectrum, microscopy allows us to see cells and their functional subunits called organelles, which can be thought of as the internal organs of a cell. We can also visualize certain proteins that may be expressed within certain organelles using fluorescence microscopy. With fluorescence microscopy, proteins in tissue or cells are tagged with light emitting markers, called fluorophores. Fluorophores make proteins under the microscope light up like fireflies on a dark summer night. Different color fluorophores can be used simultaneously to image different proteins at once, however this is limited by the number of colors available in the visible light spectrum. This means that with fluorescence imaging on a confocal microscope, there are a limited number of proteins that can be imaged within a given sample.
That’s where MARS comes in (not the planet!)! Manhattan Raman Scattering (MARS) is a special dye pallet that, combined with signals from an electronic pre-resonance Stimulated Raman Scattering microscopy (epr-SRS), creates a very sensitive way to probe, visualize and image organelles with vibrational contrast, as opposed to just light contrast. Vibrational contrast detects molecules based on their chemical properties. For example, if a probe molecule has a double bond, it will have a different vibrational frequency than a molecule with a single bond. With this technology, you can differentiate cellular targets by using dyes/probes that vary by light and vibrational signals, making these techniques very sensitive. However, these MARS dyes are difficult to chemically synthesize, and there were initially only a limited number of usable MARS dyes.
Columbia postdoc Dr. Yupeng Miao and colleagues published an article in 2021, summarizing their development of new MARS dyes that have different properties that are easier to synthesize and can visualize even more of the cell’s proteins under the microscope at once! The research contributed 30 new MARS probes that can specifically label various proteins of interest within a given sample.
Before synthesizing these new MARS probes, the researchers designed and simulated models for each potential dye. For the design, they used a similar foundation to the previous MARS probes, but included some adjustments like changing the core atom or substituting stable isotopes throughout the molecule. The results from the design models gave the researchers confidence that they could synthesize these edited molecules to expand the list of available MARS probes.
Indeed, they expanded the list of probes by developing 30 new molecules that are able to label specific cell organelles and functions. For example, MARS probes were used to image subcellular structures including the protein alpha-tubulin, which is a protein within microtubules that provide structural support to the cell, as well as fibrillarin, which is a protein that is used as a nucleoli marker. MARS probes were also shown to successfully target the cell membrane, mitochondria, lysosomes, and other lipid structures within the cell. Even more exciting – this technology allows researchers to probe each of these cellular structures simultaneously, moreso than can be done with current fluorescent microscopy methods. This means that the new MARS probes can be used to image multiple cellular markers within the same sample!
With this technology, scientists can now see even more of the unseen, which can expand our knowledge on cellular (dys)function in health and disease.
Edited by: Maaike Schilperoort, Trang Nguyen