Effectiveness of Mycobacterium tuberculosis recombinant proteins-coated gold nanoparticles in improving the interferon gamma release assay test

BACKGROUND AND OBJECTIVES In the last few decades, it has become important to find new assays to diagnose tuberculosis. Gold nanoparticles (GNPs) are chemically inert, have low toxicity, and are easy to modify and functionalize for the detection of many pathogens. They have excellent immune modulatory and adjuvant properties. In this study, GNPs were coated with recombinant Mycobacterium tuberculosis proteins, including TB۱۰.۴ (Rv۰۲۸۸), CFP-۱۰ (Rv۳۸۷۵), ESAT-۶ (Rv۳۸۷۴) and TB۷.۷ (Rv۲۶۵۴c) and exposed to the whole blood of subjects with active tuberculosis (aTB), Latent tuberculosis infection (LTBI) and healthy controls (HC). Then, the level of interferon gamma (IFN-γ) produced in the serum of the subjects was measured and compared with the level of IFN-γ produced in tubes without GNPs.MATERIALS AND METHODS The TB ۷.۷ gene was extracted and cloned from Mycobacterium tuberculosis H۳۷Rv. This antigen was expressed and purified along with other antigens: CFP-۱۰, ESAT-۶, and TB ۱۰.۴. Also, to increase the stability of recombinant proteins, different formulas prepared from additive compounds were investigated.Gold nanoparticles coated with the optimal concentration of proteins. Whole blood samples from subjects were collected in tubes containing QFT-A (CFP-۱۰, ESAT-۶, TB۷.۷, TB۱۰.۴), QFT-B (CFP-۱۰, ESAT-۶, TB۷.۷), QFT-NG (proteins coated with gold nanoparticles), TB۱ and TB۲ tubes (QFT-G-IT). Secreted interferon gamma (IFN-γ) was measured by ELISA method and results were analyzed by statistical methods.RESULTS AND DISCUSSIONThe results showed that the IFN-γ production in the GNPs tubes (QFT-NG) was significantly higher than in tubes without GNPs [QFT-A: (CFP-۱۰, ESAT-۶, TB۷.۷, TB۱۰.۴), QFT-B: (CFP-۱۰, ESAT-۶, TB۷.۷)] in aTB, LTBI, and HC subjects. The QFT-NG tube with the QFT-A tube (aTB): mean difference (MD = ۰.۴۴, ۹۵ % confidence interval: ۰.۰۷-۰.۸۸) and the QFT-NG tube with the QFT-A tube (LTBI): (MD = ۰.۲۱, ۹۵% CI: ۰.۱۵-۰.۴). Further, the group’s analysis revealed that the MD of IFN-γ production between the QFT-NG, QFT-A, and QFT-B tubes was significantly higher in aTB subjects than in LTBI subjects. Interestingly, IFN-γ production was lower in aTB and LTBI subjects in each of the TB۱ and TB۲ tubes than in the QFT-NG tube.CONCLUSIONInterferon-gamma release assays with improved sensitivity by adding gold nanoparticles increase the possibility of more accurate and faster detection of active TB and LTBI subjects.

کلیدواژه ها:

Mycobacterium Tuberculosis ، Gold Nanoparticle ، Active Tuberculosis ، Latent Tuberculosis Infection ، Interferon Gamma ، Recombinant Proteins