Antibodies play a key role in the immune system. These proteins stick to harmful substances, such as bacteria and other disease-causing pathogens, marking them for destruction or blocking their attack. Antibodies are highly selective, and this ability has been used to target particular molecules in research, diagnostics and therapies.

Typically, antibodies need to stick to a particular segment, or ‘epitope’, on the surface of a cell in order to trigger an immune response. Knowing where these regions are can help explain how these immune proteins work and aid the development of more effective drugs and diagnostic tools.

One way to identify these sites is to measure the nano-distance between antibodies and other features on the cell surface. To do this, researchers take multiple images of the cell the antibody is attached to using light microscopy. Various statistical methods are then applied to create an ‘average image’ that has a higher resolution and can therefore be used to measure the distance between these two points more accurately. While this approach works on fixed shapes, like a perfect circle, it cannot handle human cells and bacteria which are less uniform and have more complex surfaces.

Here, Kumra Ahnlide et al. have developed a new method called SiteLoc which can overcome this barrier. The method involves two fluorescent probes: one attached to a specific site on the cell’s surface, and the other to the antibody or another molecule of interest. These two probes emit different colours when imaged with a fluorescent microscope. To cope with objects that have uneven surfaces, such as cells and bacteria, the two signals are transformed to ‘follow’ the same geometrical shape. The relative distance between them is then measured using statistical methods. Using this approach, Kumra Ahnlide et al. were able to identify epitopes on a bacterium, and measure distances on the surface of human red blood cells.

The SiteLoc system could make it easier to develop antibody-based treatments and diagnostic tools. Furthermore, it could also be beneficial to the wider research community who could use it to probe other questions that require measuring nanoscale distances.

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