Absolute Quantification of Plasma Membrane Receptors Via Quantitative Flow Cytometry.

Abstract

Plasma membrane receptors are transmembrane proteins that initiate cellular response following the binding of specific ligands (e.g., growth factors, hormones, and cytokines). The abundance of plasma membrane receptors can be a diagnostic or prognostic biomarker in many human diseases. One of the best techniques for measuring plasma membrane receptors is quantitative flow cytometry (qFlow). qFlow employs fluorophore-conjugated antibodies against the receptors of interest and corresponding fluorophore-loaded calibration beads offers standardized and reproducible measurements of plasma membrane receptors. More importantly, qFlow can achieve absolute quantification of plasma membrane receptors when phycoerythrin (PE) is the fluorophore of choice. Here we describe a detailed qFlow protocol to obtain absolute receptor quantities on the basis of PE calibration. This protocol is foundational for many previous and ongoing studies in quantifying tyrosine kinase receptors and G-protein-coupled receptors with in vitro cell models and ex vivo cell samples.