Quantification of membrane-bound cytokine receptors by calibrated flow cytometry

Abstract

Summary: We present a protocol for quantifying the expression of the receptor gp130 using a calibrated flow cytometric approach. We describe pitfalls for receptor quantification such as titration of primary antibodies and standardizing cell culture. Receptors are stained with primary antibodies and fluorophore-coupled secondary antibodies. Beads covered with defined numbers of immunoglobulin G stained with fluorophore-coupled secondary antibodies serve as calibrators. In this way, the fluorescence intensity of cells is converted to the number of receptors on the cell surface.For complete details on the use and execution of this protocol, please refer to Reeh et al. (2019).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Single Cell
• Flow Cytometry/Mass Cytometry
• Signal Transduction