Fluorescence assessment of antibody binding and molecular interactions

We observed a pronounced decrease in the binding affinity of TMR to the immunoglobulin specific for this dye upon adding β-cyclodextrin. Experimental evidence suggests that TMR interacts simultaneously with the IgG antigen binding site and with the CD cavity. Fluorescence anisotropy was employed to further characterize the nature of the interactions between TMR and IgG. It is found that TMR binds with high affinity to IgG, but retains its ability to rotate within the antigen binding site.