A combinatorial dual-immunoassay predicts reactivation risk for patients with previously incomplete treatment for latent tuberculosis infection

Rationale: Diagnosis of latent TB infection (LTBI) remains challenging. The combination of QuantiFERON-TB Gold In-Tube (QFT) and flow cytometric (FC) detection of T-cell co-expression of CD25 (IL-2α receptor) and CD134 (OX40) after ex vivo TB-antigen specific challenge can visualize distinct immune subsets between unexposed subjects, untreated LTBI patients, and treated LTBI patients, which correlates with distinct TB reactivation risk predictions. We hypothesize that this strategy would identify patients with prior incomplete LTBI therapy and predict risk of TB reactivation. Objective: To apply this combinatorial dual-immunoassay analysis to subjects who were excluded from the original study, and to compare predicted cumulative risk of TB reactivation. Methods: QFT was combined with a FC assay that detects T-cell CD25 + CD134 + co-expression after TB-antigen stimulations in peripheral blood mononuclear cells. Analysis was based on previously determined technical cut-offs, and 95% bivariate normal density ellipse prediction. A modified “Online TST/IGRA interpreter” predictive formula was used. Results: We studied 74 subjects. Three out of 8 patients with history of previously incomplete LTBI therapy were identified with a combination of QFT(+) and FC assay(+) for CD4 + T-cells. Subjects with this immunoreactivity profile had the highest risk of TB reactivation (3.62± 2.60%). Conclusion: These findings further support that the strategy of combinatorial multi-immunoassay analysis is capable of generating subset profiles that can distinguish infection and treatment status, and probably confer distinctive risk of TB reactivation potential.