Calculating spectral variants when using both beads and cells as single stain controls

[Krijgsmanjens]

Hi there,

I've been wanting to try the spectral variants function in the AutoSpectral script. First of all, thanks for the great work on this!

I had a question about calculating spectral variants when using both cell-based and bead-based single-stain controls. As I understand it, the AF spectra of the single-stain controls needs to be called to calculate these variants. How should I handle this when some of my single-stain controls are cells and others are beads?

Thanks a lot in advance!

[DrCytometer]

Great question!

This is handled automatically. The AF spectra are only needed for the cell controls and are not used at all for the bead controls. The bead controls use the internal negative or assigned universal negative, without any AF extraction. You need to provide the AF spectra that best match your single-stained cell controls, which will not necessarily be the same as what you're using in the fully stained sample.

This will be handled better, and will hopefully be easier to understand in the upcoming rollout of AutoSpectral in the Honeychrome interface: https://honeychrome.cytkit.com/

[Krijgsmanjens]

Thanks for the quick answer Oliver, this clears things up a lot! I also did not know that Honeychrome was a thing, thanks for the heads-up. I'm excited to give it a shot once AutoSpectral is integrated!