Browsing by Author "Cashmore, Tamaryn J"

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    Feasibility and diagnostic utility of antigen-specific interferon-gamma responses for rapid immunodiagnosis of tuberculosis using induced sputum
    (Public Library of Science, 2010) Cashmore, Tamaryn J; Peter, Jonathan G; Van Zyl-Smit, Richard N; Semple, Patricia L; Maredza, Alice; Meldau, Richard; Zumla, Alimuddin; Nurse, Barbara; Dheda, Keertan
    BACKGROUND: The diagnosis of smear-negative or sputum-scarce tuberculosis (TB) is problematic as culture takes several weeks and representative biological samples are difficult to obtain. RD-1 antigen-specific interferon-γ release assays (IGRAs) are sensitive and specific blood-based tests for the diagnosis of M. tuberculosis infection. The feasibility and diagnostic utility of this rapid immunodiagnostic assay, using cells from induced sputum, is unknown. METHODOLOGY/PRINCIPAL FINDINGS: Cells isolated from induced sputum were co-cultured with ESAT-6 and CFP-10 antigens using a standardized enzyme-linked immunospot (ELISPOT) assay (T-SPOT®. TB ) in 101 consecutively recruited TB suspects or non-TB controls. An optimization phase using 28 samples was followed by a validation phase using samples from 73 participants (20 with definite or probable TB, and 48 with non-TB). Despite optimization of sputum processing 65/73 (89%) of the IGRAs in the validation phase were inconclusive. 44/73 (60%) tests failed due to sputum induction-related factors [sputum induction-related adverse events (n = 5), inadequate sputum volume (n = 8), non-homogenisable sputum (n = 7), and insufficient numbers of cells to perform the assay (n = 24)], whilst 20/73 (27%) tests failed due T-SPOT®. TB assay-related factors [excessive debris precluding reading of spots in the ELISPOT well (n = 6), failure of the positive control (n = 11), or high spot count in the negative control (n = 3)]. Only 8/73 (11%) of the available samples could therefore be correctly categorized (7 definite or probable TB, and 1 non-TB patient). Thus, 13/20 (65%) of the definite or probable TB cases remained undiagnosed. Conclusions/Significance: Rapid immunodiagnosis of pulmonary TB by antigen-specific IFN-γ ELISPOT responses, using cells from induced sputum, is possible. However, the test, in its current ELISPOT format, is not clinically useful because the majority of the assays are inconclusive.