Table 3.
Non-disease-specific immunological tests used for the diagnosis of PIDs
| Immunological Tests | Method(s) |
|---|---|
| Complete blood count (CBC) with differential | Automated hematology analyzer |
| Immunoglobulin quantitation - IgG, IgA, IgM, IgD, IgE | Immunoassay methods* |
| IgG, IgA subclass quantitation | Immunoassays |
| Lymphocyte subset quantitation - T, B and NK cells | Flow cytometry (FC) |
| B cell subset immunophenotyping (naïve B cells, memory B cell subsets, transitional B cells, plasmablasts, CD21low B cells) | FC |
| T cell subset immunophenotyping (T cell subsets - naïve, activated and memory, Th17 T cells, regulatory T cells) | FC |
| NK cell subset immunophenotyping (Cytotoxic and cytokine-producing NK cells, NKT cells, measurement of perforin, granzyme A, granzyme B, IFN-gamma, CD107a/CD107b for functional proteins) | FC |
| Complement pathways (classical, alternate, mannose-binding lectin) | Immunoassays, Hemolytic assays |
| Cytokines | In plasma or tissue culture, after T cell stimulation (multiplex methods - Luminex® or flow cytometry), in cells by intracellular flow cytometry, ELISPOT |
| Soluble activation or inflammatory markers - e.g. soluble BAFF, soluble CD25 (IL-2R) | Immunoassays or multiplex flow cytometry |
| Antibody responses to vaccine antigens Diphtheria, tetanus, Pneumococcal, Hemophilus influenzae among others) | Serological methods, multiplex methods (e.g. Luminex®) |
| Lymphocyte proliferation (mitogens, antigens, anti-CD3 stimulation) | Thymidine (3H-t) method, FC (CFSE, Edu®) |
| Thymopoiesis (TREC, CD4/CD8 recent thymic emigrants) | Real-time PCR, FC |
| TCR receptor diversity | Spectratyping -molecular, FC |
| NK cytotoxicity (spontaneous killing, ADCC, IL-2-stimulated and PHA stimulated cytotoxicity) | Radioactive method, FC |
| CD8 T cell cytotoxicity - mitogen-stimulated, antigen-specific | Radioactive method, FC |
| Costimulatory molecules | FC |
| TLR signaling pathways and phosphorylated proteins | FC, specific cytokines after TLR stimulation, Immunoblot analysis |
| Mutation analysis for monogenic defects of immune components | DNA-based gene sequencing |
| Measurement of innate immune responses | FC |
| Chromosomal studies for chromosomal defects - deletion, translocations and rearrangements | Fluorescence in-situ hybridization (FISH), array comparative genomic hybridization (aCGH) |
| Antigen-specific T cell quantitation | Tetramers/Pentamers/Dextramers® by FC |
| Adenosine deaminase (ADA), Purine nucleoside phosphorylase (PNP), Gluocse-6 phosphate dehydrogenase (G6PD), Myeloperoxidase (MPO) | Enzyme assays |
| Adhesion molecules for Leukocyte Adhesion deficiencies (CD18, CD11a, CD11b, CD15) | FC |
| Neutrophil oxidative burst^ | DHR test by FC (Nitroblue tetrazolium -NBT- test can also be used) |
| Delayed type Hypersensitivity | In vivo skin test |
| Autoantibodies (for PID-associated autoimmunity or autoantibody-related cytopenias) | Direct antiglobulin test (DAT or Coombs' test) for autoimmune hemolytic anemia, Immunoassays |
*For a detailed list of immunoassay methods (see Table 3, page 11, Chapter 3 - Protein Analysis for Diagnostic Applications, by AT Remaley and GL Hortin, In Molecular Clinical Laboratory Immunology, Eds, Detrick, Hamilton and Folds, 7th Ed), ^ Neutrophil chemotaxis and phagocytic cells have limited clinical utility, DHR - Dihydrorhodamine 1,2,3; a bone marrow biopsy can be performed for further evaluation of certain PIDs, e.g. abnormal retention of neutrophils in the marrow (myelokathexis in WHIM syndrome), aberrant production of hematopoietic precursors (Reticular dysgenesis and congenital neutropenias).