Interferon-gamma-release assays: Better than tuberculin skin testing?

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ABSTRACTAlthough the tuberculin skin test has long been the standard for detecting latent tuberculosis infection, it has many limitations. Interferon-gamma-release assays are gaining acceptance as an alternative. In this paper we present cases to illustrate how these new tests can be used and how to interpret the results.


  • Prior vaccination with bacille Calmette-Guérin can cause the results of skin testing to be falsely positive, but it does not affect interferon-gamma-release assays.
  • In 2005, the US Centers for Disease Control and Prevention recommended that interferon-gamma-release assays be used in all situations in which skin testing is currently used. Updated guidelines were published on June 25, 2010.
  • Successful implementation of interferon-gamma-release assay testing requires education of everyone involved—phlebotomists, laboratory personnel, occupational health workers, and clinicians.



Tuberculin skin testing, long the standard method for detecting latent tuberculosis, 1,2 has well-known limitations. Prior vaccination with bacille Calmette-Guérin (BCG) or exposure to other nontuberculous mycobacterial species can cause false-positive results. 1,3 Errors can occur in the intradermal placement and the reading of the test. The patient must return in 48 to 72 hours for an accurate reading of the test. False-negative results can occur in severe illness or immunosuppression. And a “booster response” can occur, in which immunologic memory of an earlier skin test can provoke a false-positive response. 1,3–5

Interferon-gamma-release assays are an alternative. The QuantiFERON-TB Gold test (Cellestis, Carnegie, Australia) was approved by the US Food and Drug Administration in 2001. Subsequently, two other tests were approved and are now commercially available:

  • QuantiFERON-TB Gold In-Tube (QFTGIT) (Cellestis)
  • T-SPOT.TB (Oxford Immunotec, Marlborough, MA).

We discuss how these tests work, focusing mainly on the QFT-GIT, and we present several cases to illustrate how they are used in preemployment screening and in sequential-testing surveillance programs for health care workers, and potential challenges in interpreting the results.


Unlike tuberculin skin testing, interferongamma-release assays are blood tests. 1

Either whole blood (in the QuantiFERON tests) or peripheral blood mononuclear cells (in the T-SPOT.TB test) are incubated with various tuberculosis-specific antigens. In response to the antigens, effector T cells produce interferon-gamma, which is measured quantitatively and qualitatively by either enzyme-linked immunosorbent assay (in the QuantiFERON tests) or enzymelinked immunospot assay (in the T-SPOT. TB test). 1,6,7

The kit for the QFT-GIT test, 6 which we use, contains three heparinized tubes for blood collection:

  • A control (“nil”) tube, which contains no antigens. The purpose of this tube is to determine the patient’s “baseline” level of interferon gamma.
  • A tube containing tuberculin antigens (ESAT-6, CFP-10, and TB7.7). When blood from patients who were previously exposed to Mycobacterium tuberculosis is incubated in this tube, the T cells recognizing the tuberculin antigen produce significant amounts of interferon gamma, and levels go up above that in the control tube. The level should not increase in patients not exposed to this organism.
  • A tube containing mitogen, a nonspecific stimulant of interferon gamma production. This tube represents a “positive” control.
Blood is incubated in the three tubes for 16 to 24 hours, and then the levels of interferon gamma are assayed. The tuberculin response is reported as the level of interferon gamma in the tuberculin antigen tube minus the baseline level in the nil tube. Results are reported as positive, negative, or indeterminate ( Table 1 ). (An example of an indeterminate result is a low level of interferon gamma in the tuberculin antigen tube and a low level in the mitogen tube; this combination could reflect immunosuppression or mishandling of samples.)

These tests appear to be unaffected by previous BCG vaccination, unlike tuberculin skin testing. A meta-analysis in 2008 reported a pooled specificity of 98% for the QuantiFERON tests: 99% in patients not vaccinated with BCG, and 96% in BCG-vaccinated patients. 8 The analysis also concluded that the T-SPOT.TB test appears to be more sensitive for latent tuberculosis than the QuantiFERON tests or tuberculin skin testing. 8


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