Highly-multiplexed imaging by Cyclic Immunofluorescence (CycIF) of protein levels and protein phosphorylation states in the MCF 10A breast cell line treated with EGF or four kinase inhibitors

Cyclic Immunofluorescence (CycIF) is a fluorescence imaging assay used for multiplexed measurement of protein levels and protein phosphorylation states in single cells. The MCF 10A breast cell line was plated in 96-well plates and treated with different doses of one of five perturbagens (EGF and the kinase inhibitors Lapatinib, Selumetinib, Dactolisib, and Torkinib) for 24 hr before being fixed and subjected to nine cycles of CycIF staining according to the protocol described below.