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Clinical and Diagnostic Laboratory Immunology, March 2001, p. 339-345, Vol. 8, No. 2
1071-412X/01/$04.00+0   DOI: 10.1128/CDLI.8.2.339-345.2001

Cell-Mediated Immune Response to Tuberculosis Antigens: Comparison of Skin Testing and Measurement of In Vitro Gamma Interferon Production in Whole-Blood Culture

Rohit K. Katial,1,* Joyce Hershey,1 Tejashiri Purohit-Seth,1 John T. Belisle,2 Patrick J. Brennan,2 John S. Spencer,2 and Renata J. M. Engler1

Allergy-Immunology Department, Walter Reed Army Medical Center, Washington, D.C.,1 and Department of Microbiology, Colorado State University, Fort Collins, Colorado2

Received 7 August 2000/Returned for modification 19 October 2000/Accepted 8 December 2000

Although delayed-type hypersensitivity skin testing with tuberculin purified protein derivative (PPD) is the standard for tuberculosis screening, its variability suggests the need for a more sensitive, noninvasive test. An in vitro whole-blood assay has been proposed as an alternative. Using health care worker volunteers, we confirmed the correlation between PPD skin test (PPD-ST) results (positive, induration of >15 mm) and a standardized gamma interferon (IFN-gamma ) assay, QuantiFERON-TB (Q-IFN), manufactured by CSL Biosciences in Australia, and we evaluated Mycobacterium tuberculosis culture subfractions as potential substitutes for PPD. Twenty healthy volunteers with positive PPD-ST results and 20 PPD-ST-negative controls were enrolled. Whole blood was cultured with human PPD antigens (HuPPD), Mycobacterium avium complex (MAC) PPD, phytohemagglutinin (PHA), and four M. tuberculosis culture subfractions: low-molecular-weight culture, filtrate, culture filtrate without lipoarabinomannan, soluble cell wall proteins, and cytosolic proteins, all developed from M. tuberculosis strain H37RV. Secretion of IFN-gamma (expressed as international units per milliliter) was measured by an enzyme immunoassay. The PPD or subculture fraction response as a percentage of the PHA response was used to determine positivity. Sixteen of 20 PPD-ST-positive individuals were classified as M. tuberculosis positive by Q-IFN, and 1 was classified as MAC positive. Sixteen of 20 PPD-ST-negative individuals were M. tuberculosis negative by Q-IFN, 2 were MAC positive, and 2 were M. tuberculosis positive. The tuberculosis culture subfractions stimulated IFN-gamma production in PPD-ST-positive volunteers, and significant differences could be seen between the two PPD-ST groups with all subfractions except soluble cell wall protein; however, the response was variable and no better than the Q-IFN PPD. The agreement between the Q-IFN test and the PPD-ST was good (Cohen's kappa = 0.73). The Q-IFN assay can be a useful tool in further studies of immune responses to M. tuberculosis antigens.

* Corresponding author. Mailing address: Walter Reed Army Medical Center, Allergy-Immunology Clinic, 6900 Georgia Ave., Washington, DC 20307. Phone: (202) 782-8085. Fax: (202) 782-7093. E-mail: rohit.katial{at}amedd.army.mil.

Clinical and Diagnostic Laboratory Immunology, March 2001, p. 339-345, Vol. 8, No. 2
1071-412X/01/$04.00+0   DOI: 10.1128/CDLI.8.2.339-345.2001


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