Abstract
Primary T-cell culture is an invaluable model for investigating mechanisms underlying T-cell differentiation and function in health and disease. However, different culture conditions are required for immature versus mature CD4+ T cells. Here, we provide an improved culture protocol for immature naïve mouse CD4+ T cells, including details for splenocyte isolation, naïve CD4+ T-cell purification and differentiation, and functional evaluation via flow cytometry. This protocol can also be applied for immature human CD4+ T cells. For complete details on the execution of this protocol, please refer to Wang et al. (2019).