Receptor occupancy (RO) Assays by flow cytometry are methods of assessment of binding of a therapeutic to its target receptor.  RO assays can be as simple as measuring the number of cell surface receptors bound by a biotherapeutic agent or highly complex such as measuring receptor dynamics, depending on the scientific questions being studied.  Assessment of receptor occupancy by flow cytometry can take several approaches:

  • Free Receptor Assay – Shown in the Left Panel is an illustration of free receptors that are not occupied by biotherapeutic agent detected using either a fluorescently labeled competitive antibody or biotherapeutic agent.
  • Free and Total Receptor Assay – Shown in the Middle Panel is an illustration of free and total receptor assay design which depicts the total receptors available, recognized by a noncompetitive antibody bound to a different epitope on the receptor from the one recognized by the biotherapeutic agent. This design can be used determine the ratio of free (unoccupied) sites to total sites available.
  • Direct Assessment of Bound Receptor – Shown in the Right Panel is an illustration of direct assessment of bound receptor with biotherapeutic agent, typically detected with a fluorescently labeled anti-drug antibody.

 

 

Flow cytometry can be a powerful tool for the evaluation of Receptor Occupancy (RO) to provide an assessment of the therapeutic binding to the target receptor. This can be evaluated using direct or indirect methods. Please inquire to start the scientific discussion.