Gate, analyze, and visualize FCS data

The matrix your cytometer wrote opens as a heatmap, so a value too large to be real is visible before it touches the data. Tag your single-stain controls and the spillover is measured from those tubes instead, with each channel reporting its positive-event count and its worst leak. Applying it recounts every gate on the corrected values, and each control grows a row of compensated plots you read to see whether the correction landed.

Gates are matched to each file by channel name, so a panel acquired with the detectors in a different order still lines up. Aggregate Statistics puts files down the rows and populations across the columns, bands them by the condition you assigned, and closes each band with a mean and standard deviation. Give it two conditions with at least two files in each and a Welch's t row appears, with p-values corrected for multiple comparisons and the most significant gate leading the dot plots below.

Save the gates, the plots they sit on, and the compensation matrix they were drawn under, then apply the whole strategy to the next experiment. The library checks it against your open files first and reports every gate as fine, blocked because its channel is missing, or dropped because its parent was. Templates are shared with everyone on the project and carry a discussion thread with sign-off, which is how a lab agrees on a standard panel.

The dye reference draws every fluorochrome Conspecta knows over one shared wavelength range, with its family, its laser line, and whether your lab holds a conjugate. Pick a cytometer from your own facility and the checks run on that machine's lasers and detectors, scoring a dye you have measured from your own single-stain controls ahead of the published curve. Tick rows to compare them on the spectra chart, and save the combination as a panel.