Figure 5. Analysis of the concentration dependence of ligand-receptor binding using axial lsFCS on NCI-H1703 cells stably transfected with LRP6-mCherry and immersed in CM containing DKK1-eGFP in varying concentrations.

Panels on the left and right show the data with and without background correction, respectively. (a, b) Ligand concentration dependence of the function ${\displaystyle \mathit{\Gamma }\left(C_L\right)}$ defined in Equation 5. (c, d) Fractional occupancies, F(C_L), of receptors with DKK1 ligands. Symbols: experimental data (mean ± SEM) from five axial scans of 60 s each; lines: fits with Equation 5, yielding the parameters K_D = 0.08 ± 0.01 nM and β = 0.44 ± 0.02 with background correction and K_D = 0.10 ± 0.02 nM and β = 0.31 ± 0.01 without. (e, f) Parameter ⟨γ(t)⟩ used for photobleaching correction of the autocorrelation functions. Each point represents an individual 60 s scan. Boxes mark the 25–75% range. The median is indicated by the central line, the mean as a black square; whiskers indicate the SD.