Hybridoma Screening by Antibody Capture: Flow Cytometry/FACS with Permeabilized Cells to Detect Intracellular Binding

Flow cytometry or fluorescence-activated cell sorting (FACS) can be used to identify hybridomas secreting monoclonal antibodies to internal cellular proteins, but the cells must be permeabilized before the hybridoma supernatants are applied. In using this technique, useful controls are positive and negative cell lines with primary and secondary antibodies as well as positive and negative cell lines with secondary antibody alone.