Delayed-type Hypersensitivity Workup

  • Author: Harumi Jyonouchi, MD; Chief Editor: Russell W Steele, MD   more...
 
Updated: Aug 1, 2011
 

Laboratory Studies

Characteristics of the antigens (Ags) determine the delayed-type hypersensitivity (DTH) skin test reactivity. Conjugation of the Ag to lipids facilitates the delayed-type hypersensitivity reaction. This explains the consistent response to mycobacteria in which Ag are isolated from the lipid cell wall. Size, valence, chemical composition, and dose are additional factors that are relevant to immunogenicity. Repetitive testing with the same Ag can cause an immediate immunoglobulin E (IgE)-mediated response and may diminish the delayed-type hypersensitivity skin test reactivity. High doses of Ag that induce predominant Th2 responses, such as in miliary tuberculosis, abrogate the delayed-type hypersensitivity responses by a negative feedback mechanism that suppresses Th1 responses.

By convention, the Ags used for delayed-type hypersensitivity skin testing are injected intradermally into the volar surface of the forearm with a volume of 0.1 mL each. Erythema and induration are measured at 24, 48, and 72 hours. A reaction at 24 hours does not represent DTH induced by cell-mediated immunity (CMI), or type IV reactivity. The Food and Drug Administration (FDA) –approved antigens for delayed-type hypersensitivity skin testing are limited to PPD of M tuberculosis and Candida Ag.

Conventionally, children are tested with Candida and Dermatophytin in a 1:10 or 1:100 dilution and tested with tetanus in a 1:10 or 1:100 dilution of the diphtheria-tetanus (DT) vaccine. The higher dilution is used when the child has undergone a significant infection or unusually frequent immunization respectively.

Adults are initially tested with the 1:100 concentrations of these Ags.

When interpreting delayed-type hypersensitivity skin reactivity, whether adequate exposure to the Ags has taken place prior to the procedure must be considered. A vigorous immune response to one Ag, such as in measles infection, leads to the abrogation of other delayed-type hypersensitivity responses, for example, to PPD even though the patient is also infected with tuberculosis.

Ags that are poorly immunogenic in children and in some adults include mumps (no longer on the US market) and Trichophyton. Dinitrochlorobenzene (DNCB) and dinitrofluorobenzene (DNFB) have been superseded by in vitro assessments of cell-mediated immunity because of the risk of local tissue necrosis.

When an absent delayed-type hypersensitivity reaction is noted, screening tests for a T-cell disorder should include enumeration of T and B cell subsets, and a chest radiograph to detect the thymus. Cell surface marker analysis of peripheral blood mononuclear cells (PBMCs) by flow cytometry and in vitro lymphocyte proliferation responses or production of T cell cytokins in response to mitogens (polyclonal stimulants) and recallantigens are also indicated.

Contact sensitivity to poison ivy and nickel is determined clinically; skin testing is not considered necessary. Nickel delayed-type hypersensitivity reaction can be assessed by patch skin testing.

Adverse drug reactions to antibiotics, phenytoin, and carbamazepine may involve nonimmune or immune-mediated mechanisms. The clinical setting of a reaction at 3 days or later with manifestation of a fixed rash with induration is more suspicious of involvement of a delayed-type hypersensitivity response.

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Imaging Studies

  • A chest radiograph to determine whether the thymus is present is an appropriate screening test for T-cell disorders only in the newborn; however, the thymus may involute in stressed infants in the context of overwhelming infection or severe congenital cardiac disease.
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Other Tests

When delayed-type hypersensitivity is absent and a T-cell disorder is suspected, assess in vitro lymphocyte proliferation responses against polyclonal stimulants such as mitogens (eg, phytohemagglutinins [PHA], concanavalin A [conA], pokeweed mitogen [PWM]) and specific antigens (eg, Candida, tetanus). Measurement of production of interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and interleukin (IL)-12 in response to various stimulants can also be helpful for screening severe T cell immunodeficiency as well as in mutations in IFNGR1, IFNGR2, STAT-1, IL12P40, or IL12RB1 when such mutations are suspected. Low levels of one or more of these cytokines increase the likelihood of these mutations.

In patients with severe eczema, recurrent skin abscesses, elevated IgE, and history of frequent bone fractures, assessment of IL-17 production or IL-17 expression in T-helper cells may be helpful. This is because patients with hyper IgE syndrome caused by STAT3 mutation or DOCK8 deficiency have impaired development of Th17 cells, which is a major cellular source of IL-17.[11, 12]

Cell surface markers for monocytes, T-cells (CD4, CD8, CD28, TCR α/β, TCR γ/δ), and activated T cells (CD25, HLA-DR, and CD5) are reported to be normal in IFNGR1, IL12P40,STAT1,IL12RB1, STAT3 mutations. In profound primary T-cell deficiencies such as severe combined immunodeficiency (SCID), the pattern of cell surface marker expression of lymphocyte and natural killer (NK) cells may identify the type of T-cell defect in conjunction with the clinical manifestations.

Mutational analysis for IFNGR1, IFNGR2, STAT-1,STAT-3, IL12P40, and IL12RB1 is available in commercial laboratories and/or specific research laboratories.

Additional genes that control downstream immune responses initiated by IFN-γ in the delayed-type hypersensitivity response are recognized; IFNGR2 does not bind IFN-γ but is needed for the activation of STAT-1 and its translocation to the nucleus.

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Procedures

  • When disseminated bacille Calmette-Guérin (BCG) or nontuberculosis mycobacteria (NTM) is suspected, perform biopsy of infected sites in order to examine granuloma formation and detect acid-fast mycobacteria.
  • Tissue culture to detect mycobacteria is also indicated when disseminated BCG or NTM is suspected.
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Histologic Findings

  • Granuloma formation in an intact delayed-type hypersensitivity response shows predominant infiltrates of activated macrophages and lymphocytes that can be identified as CD4+ T cells by immunohistochemical staining.
  • When NTM infection is present, multinucleated giant cells formed by fused activated macrophages are observed in the immunocompetent host.
  • In the patient with a T-cell defect, the formed granuloma lacks CD4+ T cells and these giant cells (due to ineffective macrophage activation by T cells). Instead, granulomatous lesions are characterized by infiltrate of polymorphonuclear cells, vacuolated cells, and macrophages.
  • Mycobacteria may be present in abundance but are not frequently stained, although they are isolated by culture techniques.
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Contributor Information and Disclosures
Author

Harumi Jyonouchi, MD  Associate Professor, Division of Pulmonary, Allergy/Immunology, and Infectious Diseases, Department of Pediatrics, University of Medicine and Dentistry of New Jersey-New Jersey Medical School

Harumi Jyonouchi, MD is a member of the following medical societies: American Academy of Allergy Asthma and Immunology, American Academy of Pediatrics, American Association of Immunologists, American Medical Association, Clinical Immunology Society, New York Academy of Sciences, Society for Experimental Biology and Medicine, Society for Mucosal Immunology, and Society for Pediatric Research

Disclosure: Nothing to disclose.

Specialty Editor Board

Terry W Chin, MD, PhD  Associate Director, Pediatric Allergy/Immunology/Pulmonology, Miller Children's Hospital, Long Beach Memorial Medical Center; Associate Professor, Department of Pediatrics, University of California, Irvine, School of Medicine

Terry W Chin, MD, PhD is a member of the following medical societies: American Academy of Allergy Asthma and Immunology, American Association of Immunologists, American College of Allergy, Asthma and Immunology, American College of Chest Physicians, American Thoracic Society, California Thoracic Society, Clinical Immunology Society, and Western Society for Pediatric Research

Disclosure: Nothing to disclose.

Mary L Windle, PharmD  Adjunct Associate Professor, University of Nebraska Medical Center College of Pharmacy; Editor-in-Chief, Medscape Drug Reference

Disclosure: Nothing to disclose.

John Wilson Georgitis, MD  Consulting Staff, Lafayette Allergy Services

John Wilson Georgitis, MD is a member of the following medical societies: American Academy of Allergy Asthma and Immunology, American Academy of Pediatrics, American Association for the Advancement of Science, American College of Chest Physicians, American Lung Association, American Medical Writers Association, and American Thoracic Society

Disclosure: Nothing to disclose.

David Pallares, MD  Clinical Assistant Professor, Department of Pediatrics, Division of Allergy and Immunology, University of Louisville School of Medicine

David Pallares, MD is a member of the following medical societies: American Academy of Allergy Asthma and Immunology

Disclosure: Nothing to disclose.

Chief Editor

Russell W Steele, MD  Head, Division of Pediatric Infectious Diseases, Ochsner Children's Health Center; Clinical Professor, Department of Pediatrics, Tulane University School of Medicine

Russell W Steele, MD is a member of the following medical societies: American Academy of Pediatrics, American Association of Immunologists, American Pediatric Society, American Society for Microbiology, Infectious Diseases Society of America, Louisiana State Medical Society, Pediatric Infectious Diseases Society, Society for Pediatric Research, and Southern Medical Association

Disclosure: Nothing to disclose.

Additional Contributors

The authors and editors of eMedicine gratefully acknowledge the contributions of previous author Ann O'Neill Shigeoka, MD, to the original writing and development of this article.

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