One summary number can hide where a measurement changes across the plate.
384 wells · illustrativeCols 1–2 high · 23–24 low
This map is an illustration, not experimental data.
Start with the raw wells
Inspect the spatial pattern alongside the control distributions. Look for edge patterns, row or column shifts, and unusual individual wells. Compare the map with the dispensing layout, control positions, and reading order.
Use the pattern to choose a check
Use the pattern to choose the next control. Keep preparation and reader settings with the original data so you can compare runs.
A higher Z′ can reflect nonlinear detector compression rather than a better assay. This article explains how bright-well variation can shrink while effect sizes become distorted, and how to check proportionality without confusing detector behavior with assay chemistry.
CellTiter-Glo measures ATP-dependent light, not cell number directly, so metabolism and detection conditions can change the signal independently of abundance. The article separates biological effects from reagent, temperature, mixing, and detector effects and proposes controls for each.
Six representative high-content imaging families compared by optics, simultaneous channels, live-cell conditions and the work required to turn a plate into reliable measurements.