Table 1.
Lymphocyte phenotyping, disease specific protein analysis and functional assays for IEI diagnosis.
| Test | Cell populations | Indications |
|---|---|---|
| Immunophenotyping of circulating lymphocytes | ||
| Basic T-B-NK phenotyping | CD4+ T cells, CD8+ T cells, B cells, NK cells | Basic screening for IEI, CID, SCID, PAD |
| Extended T-cell phenotyping | ||
| T1 panel | naïve (CD45RA+CCR7+), memory (CD45RO+) and CD8+ TEMRA (CD45RA+CCR7−) | SCID, OS, leaky SCID, CID, CVID |
| T2 panel | RTE (CD4+CD45RA+CD31+) | SCID, OS, leaky SCID, DiGeorge syndrome |
| T3 panel | Th1 (CXCR3+CCR6−), Th2 (CXCR3−CCR6−), Th17 (CXCR3−CCR6+), Tfh (CXCR5+) | HIES, CMC, CID |
| T4 panel | TCRαβ DNT (CD3+TCRαβ+CD4−CD8−) | ALPS, Patients with >7% of CD3+CD4-CD8- T cells |
| T5 panel | Treg (FoxP3+CD25+) | IPEX syndrome |
| Extended B-cell phenotyping | ||
| B panel | switched memory (CD27+sIgD−), non-switched memory (CD27+sIgD+), transitional B cells (CD38hiCD24hi), plasmablasts (CD38hiCD24−) and CD21lo B cells (CD38lowCD21low) | CVID, HIMS, PAD, CID, LRBA deficiency, RIPK1 deficiency |
| Analysis of the expression of specific surface or intracellular proteins | ||
| CD132 | B cells | T−B+NK- SCID |
| CD127 | T cells | T−B+NK+ SCID |
| HLA-DR | B cells | CID with low CD4 (MHC-II deficiency) |
| HLA-ABC | lymphocytes | CID with low CD8 (MHC-I deficiency) |
| ZAP-70 | T cells | CID with low CD8 |
| CD40L | PMA/Ionomycin stimulated CD4+ T cells | X-linked HIMS |
| CD40 | B cells | HIMS |
| WASp | T cells | WAS |
| Btk | monocytes | XLA |
| Perforin | NK cells | FLH 2 |
| CD18/CD11a | neutrophils | LAD-1 |
| CD15 | neutrophils | LAD-2 |
| IFN-γR1 (CD119) | monocytes | MSMD |
| IL-12Rβ1 (CD212) | PHA stimulated T cells | MSMD |
| MCP/CD46 | lymphocytes | aHUS |
| DAF/CD55 | neutrophils | PLE (CHAPLE disease) |
| Functional assays | ||
| Oxidative burst assay (DHR 123 assay) |
neutrophils | CGD, EO-IBD |
| IL-17/IFN-γ production | PMA/Ionomycin stimulated CD4+ T cells | HIES, CMC |
| STAT3/STAT1 phosphorylation assays | IL6/IFN-γ stimulated T cells/monocytes | HIES, MSMD, CMC |
| NK-cell degranulation assay | PMA/Ionomycin/Brefeldin stimulated CD3−CD56+ NK cells | FLH 3, 4 and 5, CHS, GS2 |
aHUS, atypical hemolytic uremic syndrome; ALPS, autoimmune lymphoproliferative syndrome; Btk, Bruton’s tyrosine kinase; CGD, chronic granulomatous disease; CID, combined immunodeficiency; CHAPLE, complement hyperactivation angiopathic thrombosis and protein-losing enteropathy; CHS, Chediak-Higashi syndrome; CMC, chronic mucocutaneous candidiasis; CVID, common variable immunodeficiency; DAF, decay accelerating factor; DHR, Dihydrorhodamine; EO-IBD, early onset inflammatory bowel disease; FHL, familial hemophagocytic lymphohistiocytosis; GS2, Griscelli syndrome type 2; HIMS, hyper-IgM syndrome; HIES, hyper-IgE syndrome; HLA, human leukocyte antigen; IEI, inborn errors of immunity; IPEX, immune dysregulation-polyendocrinopathy-enteropathy-x-linked; LRBA, LPS-responsive beige-like anchor protein; LAD, leukocyte adhesion deficiency; MCP, membrane cofactor protein; MHC, major histocompatibility complex; MSMD, mendelian susceptibility to mycobacterial disease, OS, Omenn syndrome, PAD, predominantly antibody deficiency; PHA, phytohemagglutinin; PLE, protein losing enteropathy; PMA, phorbol-12-myristate-13-acetate; RIPK1, receptor-interacting serine/threonine-protein kinase 1; RTE, recent thymic emigrants; SCID, severe combined immunodeficiency; sIgD, surface IgD; TEMRA, terminally differentiated effector memory CD45RA+; Treg, regulatory T cells; WAS, Wiskott Aldrich syndrome; ZAP-70, Zeta-associated protein 70.