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 |