*3.2.3 CTLA4 (CD152)*

Cytotoxic T lymphocyte Antigen-4 (CTLA4) is an inhibitor ligand of T lymphocytes which bind to CD80/CD86 which is found on antigen presenting cells with higher affinity than a costimulator molecule CD28 [8–10]. CTLA4 ceases signaling axes in T lymphocytes due to its ITIM motifs in the intracytoplasmic domain. Therefore CTLA4 blocks T cell proliferation and act important function in homeostasis and peripheral tolerance. CTLA4 is constitutively expressed on T lymphocytes and it is expressed on cell surface only after stimulation via TCR and Ca+/Calcineurin pathway *in vitro*. In patients with autosomal dominant mutation of CTLA4, lymphadenopathy/splenomegaly, hypogammaglobulinemia, cytopenia and organ specific autoimmunity are observed. This disease is also called "haploinsufficiency with autoimmune infiltration (CHAI) disease" and characterized by unfunctional or loss of CTLA4 expression on T lymphocytes [8–10]. Using flow cytometric approach, suspicious patients with CHAI disease may be investigated for molecular diagnosis before sequencing. **Figure 6** demonstrates the gating strategy for CTLA4 expression in healthy control and a patient with PID. Flow cytometry protocol for CTLA4 activation and staining are below:
