Stain index
- Sep 30, 2023
- 1 min read
The Stain Index is a measure of reagent performance on a specific cytometer, a normalized signal over background metric.
Stain Index = Brightness / Width of negative = (MFI positive – MFI negative) / 2rSD negative
∆ The brightness is a function of the assay (antigen density, fluorochrome used).
∆ The width of the negative is a function of: Instrument performance (Qr, Br, and SDen) and the assay (Fluorescence spillover / compensation), The cell population.
Corelation of Stain Index, Qr, and Br
If stain index is high then Qr is high while Br is low but at low SI it was inveresed.
Courtesy : BD Biosiences
HI There is conceptual error in the relationship between Stain Index, Qr, and Br. Actually, Stain Index measures how well a positive population is separated from the negative population, normalized to the spread (robustness) of the negative peak.
Qr (Coefficient of variation of the positive population / Resolution) represents the spread/variability of the positive population, reflects the quality of staining on the positive cells. It is related to the brightness and consistency of the fluorochrome-antibody binding. Note that
Higher Qr = more spread in the positive peak = worse resolution
Br (Background/Negative population spread) on the other side, represents the spread of the negative population. It is driven by autofluorescence, non-specific binding, and electronic noise. Remember that
Higher Br = more spread in…