T-B coculture assay for functional analysis of antigen-specific memory CD4⁺ T cells

The B cell "help" function of CD4⁺ T cells is critical in establishing the humoral arm of adaptive immunity. Here, we present a protocol to measure the "help" function of antigen-specific memory T cells using an autologous T-B coculture supplemented with monocytes. We describe steps for cell preparation, human cell sorting, coculture, and a flow cytometry-based assessment of B cell outputs. This protocol demonstrates enhanced sensitivity and proves useful in evaluating T-B collaboration in various contexts of health and disease. For complete details on the use and execution of this protocol, please refer to Ansari et al.¹.