Here, we designed and validated a single 14-color antibody combination for automated standardized and reproducible identification and monitoring of ≥89 distinct (e.g., functionally relevant) CD4+ T-cell populations in human blood, established age-related reference values, and investigated the presence of altered CD4+ T-cell subset profiles in three disease conditions—monoclonal B-cell lymphocytosis (MBL), systemic mastocytosis (SM), and common variable immunodeficiency (CVID)—setting the basis for application in routine clinical practice. Here, CD4 is linked to systemic mastocytosis.