Efficient stabilization of the Mycobacterium tuberculosis recombinant antigens for ex-vivo usages

BACKGROUND AND OBJECTIVESOne of the main goals of TB research is to identify antigens that have the ability to induce cellular and/or humoral immunity. These antigens can be used in diagnostic reagents or vaccine design. Therefore, the stability of these recombinant antigens is critical. The addition of additives to Mtb recombinant antigens during storage is critical for increasing its stability and prevention of degradation. In this study, the effect of additives in different concentrations on the degradation of Mycobacterium tuberculosis antigens include CFP-۱۰, ESAT-۶ and TB۷.۷ at different times and temperatures was investigated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).MATERIALS AND METHODSMtb antigens were cloned, and expressed in E. coli BL۲۱. Several formulations were prepared from the combination of additives. These formulas were combined with antigens in certain concentrations and antigens degradation was assessed at different times and temperatures by SDS-PAGE method. Also, the lyophilization of antigens, the lysis rate of red blood cells (RBC) and the cytotoxicity of white blood cells (WBC) in combination with each of the formulas were investigated.RESULTS AND DISCUSSIONThe results showed that additives such as sucrose and mannitol in concentrations of ۱.۵% and ۰.۵% respectively in combination with antigens for ۹ months at -۲۰ Ċ had the ability to preserve antigens compared to the control sample. Also, the comparison of the formulas showed that the presence of gelatin increases the stability of antigens at different temperatures, including ۲۵ °C. However, gelatin and glycerol increased the lysis of red blood cells in the tests. The results showed that the presence of additives include sucrose, mannitol, glycine and polyvinyl alcohol are necessary for better lyophilization of antigens.CONCLUSIONMtb recombinant antigens stabilization using chemical additives inhibits antigens degradation, leading to increased antigens stability and purification efficiency.

کلیدواژه ها:

Mycobacterium tuberculosis ، recombinant proteins ، Lyophilization ، proteins stability