: If internal markers (e.g., nuclear proteins) are the target, cells are fixed with formaldehyde and permeabilized with detergents like Triton X-100.
While IFC shares many preparatory steps with conventional flow cytometry, it requires specific optimizations to ensure high-quality imagery. Imaging Flow Cytometry: Methods and Protocols
: Cells must be in a single-cell suspension to avoid clogging the thin fluidic channels used for imaging. : If internal markers (e
: It allows for the quantification of intracellular features, such as the translocation of signals into the nucleus or the internalization of particles. Key Protocol Steps : If internal markers (e.g.
: Non-specific binding is prevented using anti-Fc antibodies. Cells are then stained with fluorophore-conjugated antibodies, viability dyes, or nuclear dyes like DAPI or Hoechst to provide spatial landmarks.