A new study shows how the single-molecule organization of receptors in a cellular context determines the function of antibodies, opening up new pathways for the development of cancer immunotherapies.

Therapeutic antibodies have transformed cancer treatment, yet the exact molecular mechanisms that drive their therapeutic effects have remained elusive. A team led by Ralf Jungmann (Chair Professor for Molecular Physics of Life at LMU and leader of the Molecular Imaging and Bionanotechnology research group at the Max Planck Institute of Biochemistry) has now taken an unprecedented look at how therapeutic antibodies reshape receptor organization at the single-molecule level, and how this affects the function of the antibodies.

Using Resolution Enhancement by Sequential Imaging (RESI) – a novel super-resolution microscopy technique capable of visualizing single proteins – the researchers directly imaged the nanoscale architecture of CD20 receptors and their interactions with widely used anti-CD20 antibodies such as Rituximab and Obinutuzumab.

“For the first time, we can visualize in intact cells how antibody-receptor complexes organize at the single-protein level,” explains Ralf Jungmann, senior author of the study, which was published recently in the journal Nature Communications. “These nanoscale patterns directly correlate with therapeutic function and provide a blueprint for rational antibody design.”

Visualizing antibody-receptor complexes at single-protein resolution

Therapeutic monoclonal antibodies act through multiple mechanisms – triggering immune cells, activating complement pathways, or directly inducing cell death. These effects depend on how antibodies bind and reorganize receptors on the cell surface. However, traditional imaging methods lacked the resolution to reveal these arrangements in their native cellular context.

The Jungmann team overcame this barrier by implementing multi-target 3D RESI imaging, a method that labels receptors and antibodies with orthogonal DNA barcodes and sequentially visualizes them at sub-nanometer precision. This allowed the team to resolve the organization of CD20 receptors and their bound antibodies directly on the cell membrane.

“We can now directly see how structural changes in antibody design translate into different receptor patterns and cellular responses,” says Isabelle Pachmayr, first author of the study. “This opens the door to structure-guided development of next-generation monoclonal antibodies.”

A new era for antibody research

Beyond CD20, this RESI technology is capable of investigating virtually any membrane receptor and therapeutic antibody with molecular resolution directly at intact cells. Because RESI visualizes whole cells with high throughput, it enables systematic analysis of antibody candidates at a resolution previously only accessible through cryo-EM, but now directly in the cellular environment and with unique molecular specificity.

Looking ahead, the team envisions combining RESI with the imaging of multiple receptors and intracellular signaling molecules to map entire therapeutic pathways. “For the first time, RESI combines the structure of nanoscale receptors with their function in a cellular context,” summarizes Jungmann. “This technology has the potential to fundamentally transform immunotherapies.”