Use this calculation for cell-culture or microscopy observations collected with one counting rule, assay gate, calibration, and unit convention. It is suited to documenting dilution, growth, labeling, geometry, or population fractions.

Parent generation MFI

A finite measured or assumed value.

Current MFI

A finite measured or assumed value.

The implementation follows this calculation rule: Infer the number of CFSE fluorescence halvings, and hence cell generations, from parent and current mean fluorescence intensities.

Generation halvings

The calculated generation halvings.

Counts and fluorescence measurements can be biased by clumping, debris, segmentation, gating, background signal, saturation, sampling fields, or unequal viability. A calculated percentage does not identify the biological mechanism behind a change. Ratios and percentages become unstable when their denominator is small, uncertain, or defined differently between samples.

Retain representative images or cytometry plots and include biological replicates. Orthogonal viability, proliferation, apoptosis, or direct counting assays can distinguish effects that a single cell metric conflates.

• CFSE Dilution Factor

  Estimate cell division count from CFSE fluorescence dilution using the base two logarithm of the initial over current intensity.

• Doubling Time (PDT)

  Calculate population doubling time as elapsed hours times ln(2), divided by ln(final count / initial count).

• S Phase Fraction

  Compute the fraction of cells in S phase relative to the total counted cell population.