Rationale: Although IFN-g release assays (IGRAs) are widely used to screen for Mycobacterium tuberculosis infection in high-income countries, published data on repeatability are limited. Objectives: To determine IGRA repeatability. Methods: The study population included consecutive patients referred to The Methodist Hospital (Houston, TX) between August 1, 2010 and July 31, 2011 for latent tuberculosis (TB) infection screening with an IGRA (QuantiFERON-TB Gold In-Tube; Cellestis, Carnegie, Australia). We performed multiple IGRA tests using leftover stimulated plasma according to a prospectively formulated quality control protocol. We analyzed agreement in interpretation of test results classified according to manufacturer-recommended criteria and repeatability of quantitative TB response. Measurements and Main Results: During the study period, 1,086 test results were obtained from 543 subjects. Per the manufacturer's cutpoint, the result of the second test was discordant from that of the first in 28 (8%) of 366 patients with valid test results, including 13 with an initial negative result and 15 with an initial positive result. Although agreement between repeat test results was high (k ¼ 0.84; 95% confidence interval, 0.79-0.90), the normal expected range of within-subject variability in TB response on retesting included differences of 6 0.60 IU/ml for all individuals (coefficient of variation, 14%), and 6 0.24 IU/ml (coefficient of variation, 27%) for individuals whose initial TB response was between 0.25 and 0.80 IU/ml. Conclusions: There is substantial variability in TB response when IGRAs are repeated using the same patient sample. IGRA results should be interpreted cautiously when TB response is near interpretation cut-points.Keywords: interferon-g release assay; QuantiFERON; repeatability; imprecision IFN-g release assays (IGRAs) are in vitro immunodiagnostic tests that measure effector T cell-mediated IFN-g response to synthetic Mycobacterium tuberculosis-specific polypeptides. The QuantiFERON-TB Gold In-Tube (QFT-GIT; Cellestis, Carnegie, Australia) is a commercially available IGRA that has been recommended as an alternative to the tuberculin skin test (TST) in targeted screening for M. tuberculosis infection (1).Although IGRAs are widely used in high-income countries and numerous studies have evaluated their diagnostic performance, there are limited data on the precision of IGRA results. Clinically, such data are essential because treatment decisions could be impacted by interpretation of results close to the threshold for a positive test, and for changes above or below this threshold when serial testing is performed (2). Data on test imprecision, including repeatability (serial testing under identical conditions) and reproducibility (serial testing under changed conditions) (3) (see Table 1), are required for CE (Conformité Européene) Marking in the European Union and for premarket approval of in vitro diagnostic tests by the US Food and Drug Administration (FDA) (4).Repeatability is unaffected by inte...