This study aimed to develop a DiaRD Mycobacterium tuberculosis interferon-gamma release assay (DiaRD MTB IGRA) and to evaluate its diagnostic performance in comparison with the QuantiFERON-TB Gold Plus (QFT-Plus) assay for the detection of MTB infection. The DiaRD MTB IGRA was developed and optimized by testing different concentrations of antigens/peptides in the TB1 and TB2 antigen tubes, as well as anti-IFN-γ antibody concentrations in the enzyme-linked immunosorbent assay (ELISA) microplates. The analytical performance of the improved assay, including sensitivity, linearity, and reproducibility, was assessed. For clinical validation, a total of 346 whole-blood samples were analyzed, comprising individuals with MTB infection (n = 80) and uninfected controls (n = 266), using both the DiaRD MTB IGRA and the QFT-Plus assay. The improved DiaRD MTB IGRA demonstrated a limit of detection (LoD) of 0.219 IU/mL within a linear dynamic range of 0-10 IU/mL. Reproducibility testing showed an overall coefficient variation (CV) of ≤11.4%. Among the 346 samples, 80 (23.1%) tested positive by QFT-Plus, including 27 from patients with active tuberculosis (TB) disease and 53 from individuals with TB infection (TBI), while 266 (76.9%) were negative. The improved DiaRD MTB IGRA demonstrated a sensitivity of 96.3% and a specificity of 96.2%. The positive predictive value (PPV), negative predictive value (NPV), and overall accuracy were 88.5%, 98.8%, and 96.2%, respectively. The overall agreement between the two assays was 93.7% (κ = 0.90; 95% CI: 0.84-0.95), with no statistically significant difference observed (p = 0.092). In conclusion, the improved DiaRD MTB IGRA showed good diagnostic performance and agreement with the comparator kit, suggesting that it may be useful for the detection of MTB infection.
Keywords: ELISA; Interferon gamma; M. tuberculosis; Tuberculosis infection.
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