The spatial variation in molecular properties within a tissue sample that affects biomarker interpretation.
Tissue heterogeneity manifests in multiple forms:
iFRETLoading... can specifically address functional heterogeneity—mapping where checkpoint engagement occurs versus where proteins are merely expressed.
Not All Areas Are the Same: Some parts of a tumor have lots of immune cells, others don't. Some areas have checkpoints engaged, others just have the proteins sitting there inactive.
A single measurement might miss the action happening elsewhere.
Heterogeneity has specific implications for functional measurement:
Expression testing: A tumor "positive" by bulk expression may have only focal positivity
Interaction testing: Checkpoint engagement may be concentrated at immune interfaces
Clinical relevance: The engaged regions may be most predictive of therapy response
Spatial FLIM-FRET preserves this information, enabling analysis of where interactions occur, not just whether they occur on average.
Why Location Matters: Checkpoint engagement might only happen where T cells meet tumor cells. Measuring the whole tumor as one number averages out this critical spatial information.
Spatial analysis keeps the location data intact.
