Binding to Nanopatterned Antigens is Dominated by the Spatial Tolerance of Antibodies

Although repetitive patterns of antigens are crucial for certain immune responses, an understanding of how antibodies bind and dynamically interact with various spatial arrangements of molecules is lacking. Hence, we introduce a new method where molecularly precise nanoscale patterns of antigens are displayed using DNA origami and immobilized in a surface plasmon resonance (SPR) setup. Using antibodies with identical antigen binding domains, we find that all subclasses and isotypes studied, bind bivalently to two antigens separated at distances ranging from 3 to 17 nm. The binding affinities of these antibodies change with the antigen distances, with a distinct preference for antigens separated by approximately 16 nm, and considerable differences in spatial tolerance exist between IgM and IgG and between low and high affinity antibodies.