Panel development and optimization in flow cytometry is a standard part of developing high complexity multi-color panels. Ideal detection and measurement of chosen cellular markers in spectral flow cytometry requires single stained reference controls that can display the authentic fluorophore spectra and that are highly similar to the autofluorescence signature of biological cell samples. In this study, a 36-color immunophenotyping panel was initially developed using polystyrene beads, however switching select fluorophores to Slingshot Bio’s cell mimic SpectraComp® and ViaComp® compensation/spectral unmixing controls improved unmixing for problematic channels. To get the best unmixing results for this panel with the most straightforward workflow, ViaComp Ultra XT™ – Amine was used as a single, universal unmixing control for both viability and antibody reference spectra.
