Figure 17.9.3.

Relationship of donor and acceptor concentrations to FRET. (A) Schematic diagram illustrating the effect of donor concentration on FRET. Oligomers of antigens (black dots) are shown randomly labeled with donor (white) and acceptor (gray) antibodies. The right diagram indicates one half the amount of donor antibody than that in the left panel. Although the donor fluorescence is expected to be lower for the right panel, the proximity of each donor to acceptor antibodies predicts that FRET efficiency should stay the same. The situation is very different for acceptor antibody concentration. (B and C) Antibody distributions are illustrated for donor (white) and acceptor (gray) antibodies on a hypothetical clustered three-antigen oligomer (black dots). Panel (B) shows the situation where the donor and acceptor antibodies bind their antigens, but do not interact with each other. Panel (C) shows the situation where the acceptor binds its antigen, but the donor antibody interacts with the acceptor antibody. In both cases, the right panel shows the consequence of reducing the acceptor concentration by one half, and the predicted effect on FRET.