The interferon gamma (IFN-γ) release assay (IGRA) is widely used as a diagnostic method for latent tuberculosis infection (LTBI). The QuantiFERON-TB Gold and QuantiFERON-TB Gold In-tube (QFT-IT) tests measure plasma IFN-γ levels using enzyme-linked immunosorbent assay (ELISA), and T-SPOT.TB counts IFN-γ-producing cells using enzyme-linked immunosorbent spot assay. IFN-γ mRNA was evaluated as an indicator of IGRA in comparison with QFT-IT IFN-γ ELISA in 46 subjects with active TB and in 73 at low risk for TB. Significant IFN-γ mRNA expression was detected from 30 min and peaked 4 h after stimulation with MTB antigens or mitogen. This was defined as the optimal time point for IFN-γ mRNA real-time polymerase chain reaction (PCR). The sensitivities of IFN-γ mRNA real-time PCR and IFN-γ ELISA were 84.8% (39/46) and 89.1% (41/46), respectively (no significant difference). Although the specificities of IFN-γ ELISA was 4.1% higher than that of IFN-γ mRNA real-time PCR (60.3% versus 56.2%), the difference was not statistically significant. The overall agreement between IFN-γ mRNA real-time PCR and IFN-γ ELISA was 79.8% (kappa = 0.475). Whilst there was no difference in the performance of IFN-γ mRNA real-time PCR and IFN-γ ELISA, IFN-γ mRNA real-time PCR was superior to IFN-γ ELISA in terms of the time required for detection of MTB infection.