Live cell
Adhered on coated glass, cross-section
An interactive walkthrough of the chemistry behind iterative indirect immunofluorescence imaging — what bonds form at each step, what the elution buffer breaks, and why the sample survives being stripped twenty times.
4i was developed in the Pelkmans lab at the University of Zurich. This explainer is an independent educational illustration of their published method. It is not affiliated with or endorsed by its authors.
Gut G, Herrmann MD, Pelkmans L. Multiplexed protein maps link subcellular organization to cellular states. Science. 2018;361(6401):eaar7042. doi:10.1126/science.aar7042
Kramer BA, Sarabia del Castillo J, Pelkmans L, Gut G. Iterative Indirect Immunofluorescence Imaging (4i) on Adherent Cells and Tissue Sections. Bio-protocol. 2023;13(13):e4712. doi:10.21769/BioProtoc.4712
Bailey LE. 4i, explained interactively. 2026.
laelise.github.io/4i-explainer
Please cite the papers above for the method itself.
Every scene is a schematic of interaction classes, not a structural model. Bond geometry, antibody pose and protein shape are illustrative. Buffer concentrations shown on the elution controls reflect the published working values. Nothing here contains experimental data.
Diagrams built as SVG. Use Export SVG to download the current frame as an editable vector file.