Determination of the Dissociation Constant of the NFκB p50/p65 Heterodimer in Living Cells Using Fluorescence Cross-Correlation Spectroscopy

Fluorescence cross-correlation spectroscopy (FCCS) is a promising technique for observing and quantifying protein-protein interactions in vitro and in vivo. FCCS has emerged as a useful tool for obtaining parameters of the concentration of labeled particles, their molecular dynamics, as well as the size of their complexes. This chapter discusses aspects of preparing a biological system for FCCS experiments and suggests practical advice for performing FCCS in living cells. Moreover, we describe the method of FCCS to determine the dissociation constant of a transcription factor dimer in the living cell.