Functional immune responses require the appropriate activation of antigen-specific B cells from the naïve repertoire. B cell activation is initiated when the B cell receptor (BCR) binds antigens that are displayed on the surfaces of antigen-presenting cells. This binding event triggers the B cell to internalise, process, and present the antigen to helper T cells, which provide signals required for B cell differentiation and the production of antibodies. The extent of T cell help depends upon the affinity of the BCR for antigen, suggesting that affinity discrimination during antigen acquisition is essential for high-affinity antibody responses. Despite the importance of B cell antigen internalisation for this process, the mechanisms underlying how B cells acquire antigens from the surfaces of other cells remain poorly understood. One limiting factor in studies to date has been the artificial substrates used to mimic antigen-presenting cell membranes. To overcome this limitation, we have developed DNA-based nanosensors that enable us to investigate mechanisms of B cell antigen acquisition and processing from both artificial substrates and live antigen-presenting cells. Our results show that B cell antigen internalisation is regulated by several mechanical signals including BCR affinity for antigen, the strength of the antigen tether, and the flexibility of the membrane substrate. We demonstrate that by tuning these properties we can change the mechanism by which B cells internalise antigen, the amount of antigen that B cells acquire, and the ability of B cells to discriminate low- and high-affinity antigens. These observations highlight a potential role for mechanical forces in the activation and regulation of B cell responses.