What the plate reader gives you
A plate reader exports a matrix of numbers, one per well. Depending on your instrument and assay, those numbers are:
| Format | Unit | Common assay |
|---|---|---|
| Absorbance | OD (optical density, unitless) | ELISA, protein quantification |
| Fluorescence intensity | RFU (relative fluorescence units) | Cell viability, reporter assays |
| Luminescence | RLU (relative light units) | ATP assays, luciferase reporters |
| Time-resolved fluorescence | Counts or ratio | HTRF, TR-FRET |
All of these are relative measurements. They only mean something in the context of controls on the same plate.
Validating the raw file
Before importing, open the exported file and check for these common problems:
Merged or missing headers
Plate reader software often exports files with merged cells, multi-row headers, or instrument metadata above the data block. Platelet handles most common formats automatically, but verifying manually the first time saves debugging later.
Saturated wells
A well reading at or very near the instrument's maximum value is saturated: the detector cannot distinguish between a strong signal and an overwhelming one. Typical saturation thresholds:
| Format | Saturation indicator |
|---|---|
| Absorbance | OD > 3.5 (most readers) |
| Fluorescence | Instrument max, often 65535 counts |
| Luminescence | Overflow flag in exported file |
Saturated wells should be excluded before any downstream analysis.
A saturated positive control makes your Z′-factor calculation meaningless. If your positive control is saturated, dilute it and re-run rather than proceeding with normalisation.
Checking the signal window
The signal window is the ratio of your positive to negative control means. A window below 3× is generally too small to detect weak hits reliably.
Check your signal window plate-by-plate, not as a global average across a run. A single plate with a collapsed window should be flagged and re-run, not silently averaged away.
Plate-to-plate consistency
If you are running multiple plates in a single experiment, compare the negative control means across plates before merging the data. A drift of more than 20% between plates indicates a systematic problem, such as reagent degradation, incubation time differences, or reader lamp variation.
Importing into Platelet
Platelet accepts Excel and CSV exports from all major plate reader software. Upload the file in the Readings tab, map the data block to the correct wells, and the app will flag saturated values and compute the signal window automatically before you proceed to analysis.