Laboratory diagnostic tests for example IEIs with AICs
| Disease example . | Genetic defect . | Basic laboratory tests* . | Immunophenotyping* . | Functional tests* . | Other tests/genetic testing . |
|---|---|---|---|---|---|
| CHAI | CTLA4 | CBC, serum immunoglobulins, MCV, reticulocyte count, haptoglobin, direct Coombs test, bilirubin, LDH, blood smear examination, neutrophil and platelet autoantibodies† | TBNK (C), T-cell subset quantitation (for naïve, memory effector, central, effector memory expressing CD45RA, activated T cells expressing HLA-DR, CD25, CD38, CD69, senescent/exhausted T cells negative for CD28, expressing CD57, activated/memory T cells expressing PD-1); follicular T helper cells, which are expanded; Tregs (expressing FOXP3, CD127), which are decreased; B-cell subset quantitation (naïve, transitional (CD24, CD38); memory B cells, including total CD27+, marginal zone and switched memory B cells, CD10+ immature B cells, CD21−, CD21dim, CD21++ B cells, plasmablasts, BAFF-R/TACI-expressing total and memory B cells, NK cells (cytokine-producing, CD56++, cytotoxic NK cells (CD16++56+/−) (C) | If recurrent infections are present, T-cell function can be assessed by proliferation to mitogens (PHA, anti-CD3/anti-CD28/IL-2), antigens (CA, TT, viral peptides, as needed) (C). | Expression of CTLA-4 on activated T cells and Tregs (after stimulation with PHA or PMA + ionomycin) (R), transendocytosis assay (R) if it can be assessed |
| T-cell proliferation to other mitogenic stimulants, such as PMA/ionomycin, is typically not available clinically but is useful for assessing the distal T-cell activation pathway (R). | sBAFF, sCD25 (significantly elevated), and other biomarkers if relevant to clinical phenotype | ||||
| Production of cytokines (IL-2, TNF-α, and IFN-γ) may also be assessed in T cells after stimulation with mitogens and CD4+ and CD8+ T cells (R) | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| LATAIE | LRBA | Same as for CTLA-4 defects | Same as for CTLA-4 defects | Same as for CTLA-4 defects | LRBA protein expression in lymphocyte subsets, monocytes by flow cytometry (R) |
| Transendocytosis assay (R) | |||||
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| APDS1/2, PASLI | PIK3CD (GOF), PIK3CD (LOF), PIK3R1 | Same as for CTLA-4 defects, increased serum IgM levels | Same as for CTLA-4 defects; these patients have decreased naïve T cells, increased senescent T cells, and transitional B cells. | Same as for CTLA-4 defects | Akt phosphorylation by flow cytometry or western blot (R) Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) |
| ALPS | FAS, FASLG, CASP10, others | Same as for CTLA-4 defects, vitamin B12, sFASL, which are increased; IL-10 | Quantitation of TCRαβ+ DNT (CD3+CD4−CD8−) T cells, which are increased; CD25, HLA-DR (increased ratio of HLA-DR to CD25), CD57 on T cells (C); switched memory B cells, which are decreased in some patients; γδ TCR+ T cells (C) | As needed | In vitro assessment of apoptosis for patients not yet started on treatment (C) |
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| HLH | PRF1, UNC13D, STX11, STXBP2, others | CBC, sCD25, ferritin, bone marrow biopsy, liver enzymes, IL-18, CXCL9 (surrogate for IFN-γ levels) (C) | TBNK, NK cell subset quantitation (C) | CD107 degranulation in CD8+ T cells and NK cells after PMA/ionomycin and K562 stimulation, respectively, by flow cytometry (C), which is normal in perforin deficiency and abnormal in other genetic types of HLH | Perforin protein expression in NK cells and CD8+ T cells by flow cytometry (C), which is abnormal in FHL2 but normal in other genetic types of HLH |
| Lymphopenia can be observed; decreased cytotoxic NK cells | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| GATA2 haploinsufficiency | GATA2 | CBC, bone marrow biopsy to look for atypical megakaryocytes | TBNK, monocyte subsets (for evaluation of monocytopenia); dendritic cell subsets (CD123, CD141, CD1c, CD11c), which are all significantly decreased (R); NK cell subsets for loss of CD56++ cytokine-producing NK cells (C) | T-cell function as needed (as described for CTLA-4 defects) | If concern for somatic GATA2 variant, obtain sample from sources other than blood (buccal and fibroblasts), check for presence of somatic ASXL1 variant |
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| DC | DKC1, RTEL1, TERC, TERT, others | CBC, serum immunoglobulins | TBNK, additional phenotyping as needed, assessment of DNA repair defects by flow cytometry (phosphorylation of ATM and H2AX-γH2AX)# (C) | T-cell function as needed (as described for CTLA-4 defects) | Telomere length analysis by flow cytometry–fluorescence in situ hybridization to estimate how short telomeres are in each subset: lymphocytes and granulocytes, assessment for IPF, depending on age of patient |
| Lymphopenia can be observed in one or more subsets | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| DNA repair defects are observed in some patients and can be assessed by flow cytometry‡ (C) | |||||
| WHIM | CXCR4 (GOF) | CBC (ANC often severe <500/mm3, though could be higher at times); serum immunoglobulins because some patients have hypogammaglobulinemia; BM evaluation for myelokathexis | TBNK, other phenotyping as needed | T-cell function as needed (as described for CTLA-4 defects) | Expression of CXCR4 on activated T cells, chemotaxis in response to SDF-1 (R) |
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| XLP-2 | XIAP/BIRC4 | Same as HLH | TBNK, other phenotyping as needed | T-cell function as needed (as described for CTLA-4 defects) | XIAP protein expression in lymphocyte subsets by flow cytometry§ (C) |
| CD107 degranulation; NK cell cytotoxicity can appear decreased due to NK cell lymphopenia | XIAP functional analysis|| (C) | ||||
| Normal Fas-mediated T-cell apoptosis (not increased) | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| WAS | WAS | Same as for CTLA-4 defects, platelet size measurement | As described for CLTA-4 defects, for patients with WAS who have malignancies; early data suggest DNA repair defects, which can be assessed by flow cytometry‡ (C) | T-cell proliferation to mitogens and antigens, NK cell cytotoxicity (abnormal with K562 stimulation, but normal with IL-2 (C) or IL-15 stimulation (R) | WAS protein expression by flow cytometry (C) |
| Assessment of immunological synapse formation (R) | |||||
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| STAT3-GOF | STAT3 | Same as for CTLA-4 defects, as needed | Same as for CTLA-4 defects, as needed | Same as for CTLA-4 defects, as needed | STAT1/5 phosphorylation by flow cytometry, which is decreased (C, R) |
| T, B, and NK cell lymphopenia in some patients | Increased SOCS3 expression (R), IL-6 levels (C) | ||||
| Decreased switched memory B cells | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| Variable numbers of Th17 cells | |||||
| STK4 deficiency | MST1 | Same as for CLTA-4 defects, serum IgE quantitation | Same as for CTLA-4 defects (phenotypic overlap with DOCK8 deficiency in some patients) | Same as for CTLA-4 defects | Western blot analysis for STK4/MST1 protein expression (R) |
| CD4+ T-cell lymphopenia | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| Marginal zone B cells (nonswitched) are decreased | |||||
| Switched memory B cells are normal to high |
| Disease example . | Genetic defect . | Basic laboratory tests* . | Immunophenotyping* . | Functional tests* . | Other tests/genetic testing . |
|---|---|---|---|---|---|
| CHAI | CTLA4 | CBC, serum immunoglobulins, MCV, reticulocyte count, haptoglobin, direct Coombs test, bilirubin, LDH, blood smear examination, neutrophil and platelet autoantibodies† | TBNK (C), T-cell subset quantitation (for naïve, memory effector, central, effector memory expressing CD45RA, activated T cells expressing HLA-DR, CD25, CD38, CD69, senescent/exhausted T cells negative for CD28, expressing CD57, activated/memory T cells expressing PD-1); follicular T helper cells, which are expanded; Tregs (expressing FOXP3, CD127), which are decreased; B-cell subset quantitation (naïve, transitional (CD24, CD38); memory B cells, including total CD27+, marginal zone and switched memory B cells, CD10+ immature B cells, CD21−, CD21dim, CD21++ B cells, plasmablasts, BAFF-R/TACI-expressing total and memory B cells, NK cells (cytokine-producing, CD56++, cytotoxic NK cells (CD16++56+/−) (C) | If recurrent infections are present, T-cell function can be assessed by proliferation to mitogens (PHA, anti-CD3/anti-CD28/IL-2), antigens (CA, TT, viral peptides, as needed) (C). | Expression of CTLA-4 on activated T cells and Tregs (after stimulation with PHA or PMA + ionomycin) (R), transendocytosis assay (R) if it can be assessed |
| T-cell proliferation to other mitogenic stimulants, such as PMA/ionomycin, is typically not available clinically but is useful for assessing the distal T-cell activation pathway (R). | sBAFF, sCD25 (significantly elevated), and other biomarkers if relevant to clinical phenotype | ||||
| Production of cytokines (IL-2, TNF-α, and IFN-γ) may also be assessed in T cells after stimulation with mitogens and CD4+ and CD8+ T cells (R) | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| LATAIE | LRBA | Same as for CTLA-4 defects | Same as for CTLA-4 defects | Same as for CTLA-4 defects | LRBA protein expression in lymphocyte subsets, monocytes by flow cytometry (R) |
| Transendocytosis assay (R) | |||||
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| APDS1/2, PASLI | PIK3CD (GOF), PIK3CD (LOF), PIK3R1 | Same as for CTLA-4 defects, increased serum IgM levels | Same as for CTLA-4 defects; these patients have decreased naïve T cells, increased senescent T cells, and transitional B cells. | Same as for CTLA-4 defects | Akt phosphorylation by flow cytometry or western blot (R) Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) |
| ALPS | FAS, FASLG, CASP10, others | Same as for CTLA-4 defects, vitamin B12, sFASL, which are increased; IL-10 | Quantitation of TCRαβ+ DNT (CD3+CD4−CD8−) T cells, which are increased; CD25, HLA-DR (increased ratio of HLA-DR to CD25), CD57 on T cells (C); switched memory B cells, which are decreased in some patients; γδ TCR+ T cells (C) | As needed | In vitro assessment of apoptosis for patients not yet started on treatment (C) |
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| HLH | PRF1, UNC13D, STX11, STXBP2, others | CBC, sCD25, ferritin, bone marrow biopsy, liver enzymes, IL-18, CXCL9 (surrogate for IFN-γ levels) (C) | TBNK, NK cell subset quantitation (C) | CD107 degranulation in CD8+ T cells and NK cells after PMA/ionomycin and K562 stimulation, respectively, by flow cytometry (C), which is normal in perforin deficiency and abnormal in other genetic types of HLH | Perforin protein expression in NK cells and CD8+ T cells by flow cytometry (C), which is abnormal in FHL2 but normal in other genetic types of HLH |
| Lymphopenia can be observed; decreased cytotoxic NK cells | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| GATA2 haploinsufficiency | GATA2 | CBC, bone marrow biopsy to look for atypical megakaryocytes | TBNK, monocyte subsets (for evaluation of monocytopenia); dendritic cell subsets (CD123, CD141, CD1c, CD11c), which are all significantly decreased (R); NK cell subsets for loss of CD56++ cytokine-producing NK cells (C) | T-cell function as needed (as described for CTLA-4 defects) | If concern for somatic GATA2 variant, obtain sample from sources other than blood (buccal and fibroblasts), check for presence of somatic ASXL1 variant |
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| DC | DKC1, RTEL1, TERC, TERT, others | CBC, serum immunoglobulins | TBNK, additional phenotyping as needed, assessment of DNA repair defects by flow cytometry (phosphorylation of ATM and H2AX-γH2AX)# (C) | T-cell function as needed (as described for CTLA-4 defects) | Telomere length analysis by flow cytometry–fluorescence in situ hybridization to estimate how short telomeres are in each subset: lymphocytes and granulocytes, assessment for IPF, depending on age of patient |
| Lymphopenia can be observed in one or more subsets | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| DNA repair defects are observed in some patients and can be assessed by flow cytometry‡ (C) | |||||
| WHIM | CXCR4 (GOF) | CBC (ANC often severe <500/mm3, though could be higher at times); serum immunoglobulins because some patients have hypogammaglobulinemia; BM evaluation for myelokathexis | TBNK, other phenotyping as needed | T-cell function as needed (as described for CTLA-4 defects) | Expression of CXCR4 on activated T cells, chemotaxis in response to SDF-1 (R) |
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| XLP-2 | XIAP/BIRC4 | Same as HLH | TBNK, other phenotyping as needed | T-cell function as needed (as described for CTLA-4 defects) | XIAP protein expression in lymphocyte subsets by flow cytometry§ (C) |
| CD107 degranulation; NK cell cytotoxicity can appear decreased due to NK cell lymphopenia | XIAP functional analysis|| (C) | ||||
| Normal Fas-mediated T-cell apoptosis (not increased) | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| WAS | WAS | Same as for CTLA-4 defects, platelet size measurement | As described for CLTA-4 defects, for patients with WAS who have malignancies; early data suggest DNA repair defects, which can be assessed by flow cytometry‡ (C) | T-cell proliferation to mitogens and antigens, NK cell cytotoxicity (abnormal with K562 stimulation, but normal with IL-2 (C) or IL-15 stimulation (R) | WAS protein expression by flow cytometry (C) |
| Assessment of immunological synapse formation (R) | |||||
| Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | |||||
| STAT3-GOF | STAT3 | Same as for CTLA-4 defects, as needed | Same as for CTLA-4 defects, as needed | Same as for CTLA-4 defects, as needed | STAT1/5 phosphorylation by flow cytometry, which is decreased (C, R) |
| T, B, and NK cell lymphopenia in some patients | Increased SOCS3 expression (R), IL-6 levels (C) | ||||
| Decreased switched memory B cells | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| Variable numbers of Th17 cells | |||||
| STK4 deficiency | MST1 | Same as for CLTA-4 defects, serum IgE quantitation | Same as for CTLA-4 defects (phenotypic overlap with DOCK8 deficiency in some patients) | Same as for CTLA-4 defects | Western blot analysis for STK4/MST1 protein expression (R) |
| CD4+ T-cell lymphopenia | Genomic analysis (either targeted, WES, WGS, or chromosomal array may also be helpful for an initial screen of large deletions, CNVs) (C) | ||||
| Marginal zone B cells (nonswitched) are decreased | |||||
| Switched memory B cells are normal to high |
ANC, absolute neutrophil count; APDS, activated phosphatidylinositol 3-kinase δ syndromes 1 and 2; BAFF, B-cell activating factor; BAFF-R, B-cell activating factor receptor; BM, bone marrow; C, clinically available; CA, Candida; CBC, complete blood count; CNV, copy number variation; DC, dyskeratosis congenita; DNT, double-negative (CD3+CD4−CD8−) T cells; GATA2, GATA2 haploinsufficiency; GOF, gain of function; HLH, hemophagocytic lymphohistiocytosis; IFN, interferon; IPF, idiopathic pulmonary fibrosis; LATAIE, LRBA deficiency with autoantibodies, Treg defects, autoimmune infiltration, and enteropathy; LDH, lactate dehydrogenase; LOF, loss of function; MCV, mean corpuscular volume; PASLI, p110-δ–activating mutation causing senescent T cell, lymphadenopathy, immunodeficiency; PHA, phytohemagglutinin; PMA, phorbol myristate acetate; R, research testing only; SDF-1, stromal cell–derived factor 1 (CXCL12); sFASL, soluble FAS ligand; STK4/MST1, serine threonine kinase 4, macrophage-stimulating factor 1; TBNK, lymphocyte subset quantitation (T, B, and NK cells); TCR, T-cell receptor; TNF, tumor necrosis factor; TT, tetanus toxoid; WAS, Wiskott-Aldrich syndrome; WHIM, warts, hypogammaglobulinemia, immunodeficiency, myelokathexis; XIAP, X-linked inhibitor of apoptosis; XLP-2, X-linked lymphoproliferative disease type 2.
Testing to be ordered as appropriate on the basis of clinical phenotype
†Antiplatelet and antineutrophil antibody tests have limited sensitivity and therefore are not always diagnostically valuable.
‡Cousin et al.36
Gifford et al.63
Ammann et al.64