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Engineering mycobacteria for the production of self-assembling biopolyesters displaying mycobacterial antigens for use as a tuberculosis vaccine

Applied and Environmental Microbiology. 2017; 
Jason W. Lee, Natalie A. Parlane, Bernd H. A. Rehm, Bryce M. Buddle, and Axel Heiser
Products/Services Used Details Operation
Protein bands separated by SDS-PAGE was transferred to nitrocellulose membrane using i-BLOT system (Invitrogen, Carlsbad, CA). Membrane was blocked with 1% skim milk in PBST for 1 h. Following washing with PBST, primary antibodies were diluted in 1% BSA and used accordingly: for detection of PhaC, 1:20,000 rabbit polyclonal (GenScript, NJ); ESAT-6, 0.1 μg/mL rabbit polyclonal (Abcam, Cambridge, United Kingdom); and GFP, 0.75 μg/mL rabbit polyclonal (A01388, GenScript, NJ).

摘要

Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world’s biggest global health burdens. Recently, engineered polyhydroxyalkanoate (PHA) biobeads produced in both E. coli and Lactococcus lactis displaying mycobacterial antigens were found to induce significant cell mediated immune responses in mice. We observed that such PHA beads contained host cell proteins as impurities which we hypothesized to have the potential to induce immunity. In this study we aimed to develop PHA beads produced in mycobacteria (mycobacterial PHA biobeads, MBB) and test their potential as TB vaccine in a mouse model. As a model organism, nonpathogenic Mycob... More

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