Antibody Internalization Assay Using a pH Sensor Fluorescent Dye

Therapeutic antibodies conjugated to a toxic small molecule are increasingly being developed.  A key requirement for these antibody drug conjugates (ADCs) is the ability of the antibody to bind to the antigens, typically receptors, on tumor cells and become internalized.  Binding the antigen does not necessarily mean the antibody will become internalized and appears to be epitope specific. Hence, methods that allow selection of antibodies based on ability to be internalized is an important assay early in the hybridoma screening stage.

In this webinar, you will learn about a fluorescent dye that:

• Fluoresces brightly at low pH but is non-fluorescent at neutral pH
• Conjugates easily to candidate antibodies with amine- or thiol-reactive chemistry either in colution or while immobilized on protein A or protein G beads
• Produces a high signal-to-background ratio suitable for plate-based assays as well as fluorescent microsopy and flow cytometry analysis

The utility of the pH sensitive dye will be demonstrated with examples of internalization assays using antibodies like Transtuzumab, Cetuximab and Panitumumab.