Flow Cytometry Assay Differentiates Active from Latent Tuberculosis

A flow cytometry-based assay identifies parameters of the immune response correlated to protection against tuberculosis (TB).

The assay derives functional signatures of immune responses to Mycobacterium tuberculosis (Mtb), the causative organism of TB, such as the cytokines and leucocytes, at different stages of the disease.

The new tool has been developed by scientists at the University of Lausanne, (Lausanne, Switzerland), that distinguishes between active and latent TB. A study recruited 283 participants from a hospital in Switzerland based on positive Mtb-specific interferon gamma (IFN-γ) enzyme-linked immunosorbent spot assay (Becton Dickinson, Allschwil, Switzerland). A validation cohort of 114 participants was obtained from two clinical sites from Switzerland and South Africa. For the flow cytometry analysis cells were stained with antibodies specific for a range of cytokines.

The scientists were able to show that the immune response by the cytokines such as tumor necrosis factor alpha (TNF-α), interleukin 2 (IL-2) and IFN-γ, were correlated to the different stages of tuberculosis disease. The flow cytometry-based assay read-out is focused on the immune response and not on microbiological parameters, thus explaining its reliability and quickness. The sensitivity of the assay was 62% and specificity was 92% for the different stages of TB. The results showed substantial increase in the proportion of single-positive TNF-α Mtb-specific CD4+ T cells, a type of white cell, in subjects with active disease. This parameter was the strongest predictor of diagnosis of active disease versus latent infection.

Giuseppe Pantaleo, MD, professor of medicine and lead author of the study, said, "This assay and the functional signatures of TB-specific immune responses are not only relevant for discriminating between active TB disease and latent infection, but may be also instrumental in monitoring the response to TB therapy." The authors concluded that although the flow cytometry-based assay requires specific equipment and skillful laboratory personnel, further technological development is already under way to make the assay widely accessible. The study was published online on February 20, 2011, in Nature Medicine.

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