Colocalization analysis has a reputation problem. For years, the standard practice was to merge the red and green channels, look for yellow, and declare colocalization. This… See the analysis in action: https://youtube.com/watch?v=uf0FLNjSNJ8
Colocalization analysis has a reputation problem. For years, the standard practice was to merge the red and green channels, look for yellow, and declare colocalization. This approach is subjective, sensitive to image brightness, and routinely produces misleading results. Reviewers know it. Editors know it. And yet it persists, because doing it properly, computing Pearson and Manders coefficients per cell, requires tools that most bench scientists don't have set up.
Cytely computes Pearson correlation or Manders M1/M2 overlap coefficients within each cell, not per image. This matters enormously: a per-image coefficient averages over all cells, hiding the fact that some cells show strong colocalization and others show none. Per-cell coefficients give you a distribution, the real biology.
And if you need more specificity, you can further restrict the analysis to a specific compartment (nucleus-only or cytoplasm-only).
Target engagement, proving that a drug reaches its intracellular target, often relies on colocalization of a labeled compound with a target marker. Subjective "yellow merge" images don't survive regulatory scrutiny. Per-cell Pearson coefficients with image-linked verification do.
How the assay runs
Segment cells → compute Pearson correlation and Manders coefficients between Ch2 and Ch3 within each cell. Optionally restrict analysis to a specific compartment (nucleus-only or cytoplasm-only).
What you provide
- Ch1: DAPI
- Ch2: marker A (IF or fluorescent tag)
- Ch3: marker B (IF or fluorescent tag)
What you get
- Pearson correlation coefficient per cell
- Manders M1/M2 overlap coefficients per cell
- % cells with positive colocalization
- Channel intensity ratios per cell
Ready to run
Receptor-ligand co-localization (e.g., EGFR + EGF), organelle contact sites (e.g., mitochondria-ER via MitoTracker + ER-Tracker), pathway co-activation (e.g., phospho-protein + total protein), drug accumulation localization (e.g., compound + lysosomal marker), compartment-specific colocalization (nuclear-only, cytoplasmic-only).
Inquire for support
Object-based colocalization (spot-to-spot spatial overlap scoring), cross-correlation function analysis, 3D colocalization.