Measurement of Anti-IFN-γ Autoantibodies

Measurement of anti-IFN-γ autoantibodies is conducted at Kumamoto University.

Anti-IFN-γ antibodies can be detected in patients with tuberculosis and pulmonary nontuberculous mycobacterial (NTM) disease; however, these antibodies lack neutralizing capacity ⁽¹⁾. In adult-onset immunodeficiency (AOID), anti-interferon (IFN)-γ antibodies possess neutralizing capacity. In Japan, a measurement system has been developed for the relative quantification of anti-IFN-γ antibodies using enzyme-linked immunosorbent assay (ELISA) and quantification of the neutralizing capacity through signal transducer and activation of transcription 1 (STAT1) phosphorylation ⁽²⁾.

Figure:Relative quantification of anti-IFN-γ antibodies(ELISA) — Serum or plasma was added to plates coated with human IFN-γ and ELISA was performed using anti-human IgG secondary antibodies to measure anti-IFN-γ antibodies.

Figure:Anti-IFN-γ antibodies titers in disseminated NTM cases and healthy individuals — Compared to healthy individuals and disseminated NTM cases negative for anti-IFN-γ antibodies, significant increases in anti-IFN-γ antibody titers were observed in patients with disseminated NTM patients positive for anti-IFN-γ antibodies.

Figure:IFN-γ Signaling — IFN-γ induces phosphorylation of STAT-1, a downstream transcription factor of intracellular signal transduction via IFN-γ receptors on the surface of leukocytes.
Phosphorylated STAT-1 translocates to the nucleus and regulates gene expression as a transcription factor.

Figure:Phosphorylation of STAT1 in whole blood — Phosphorylation of STAT1 after the stimulation of leukocytes in whole blood with different concentrations of IFN-γ was detected by flow cytometry. In healthy individuals, IFN-γ increased STAT1 phosphorylation in leukocytes in a dose-dependent manner. In contrast, in the presence of neutralizing anti-IFN-γ autoantibodies in the blood, STAT1 phosphorylation did not occur in leukocytes even with increased concentrations of added IFN-γ.

Figure:Evaluation of neutralizing capacity using preserved serum and plasma — In healthy individuals (●), STAT1 phosphorylation intensity increased with rising concentrations; however, phosphorylation was not detected in disseminated NTM cases positive for anti-IFN-γ antibodies (○). Comparisons among patients with disseminated NTM (dNTM), healthy controls (HC), those with pulmonary NTM (pNTM) disease, and those with pulmonary tuberculosis (pTBC) demonstrated that phosphorylation was not detected in patients with dNTM disease, thus maintaining the IFN-γ neutralizing capacity.

Reference List

Aoki A, Sakagami T, Yoshizawa K, Shima K, Toyama M, Tanabe Y, et al. Clinical Significance of Interferon-gamma Neutralizing Autoantibodies Against Disseminated Nontuberculous Mycobacterial Disease. Clin Infect Dis. 2018;66(8):1239-45.
Shima K, Sakagami T, Tanabe Y, Aoki N, Moro H, Koya T, et al. Novel assay to detect increased level of neutralizing anti-interferon gamma autoantibodies in non-tuberculous mycobacterial patients. J Infect Chemother. 2014;20(1):52-6.