TY - JOUR ID - 8363 TI - Selection of single-chain variable fragments specific for Mycobacterium tuberculosis ESAT-6 antigen using ribosome display JO - Iranian Journal of Basic Medical Sciences JA - IJBMS LA - en SN - 2008-3866 AU - Ahangarzadeh, Shahrzad AU - Bandehpour, Mojgan AU - Kazemi, Bahram AD - Department of Biotechnology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran AD - Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran Y1 - 2017 PY - 2017 VL - 20 IS - 3 SP - 327 EP - 333 KW - antibody KW - ESAT-6 KW - Mycobacterium tuberculosis KW - Ribosome display KW - scFv DO - 10.22038/ijbms.2017.8363 N2 - Objective(s): Tuberculosis (TB) is still one of the problematic infectious diseases in developing countries, especially in Iran. In the present study, we applied ribosome display technique to select single chain variable fragments (scFvs) specific for the 6-kDa early secretory antigenic target (ESAT-6) antigen of Mycobacterium tuberculosis from a mouse scFv library. Materials and Methods: The gene encoding ESAT-6 was cloned into pET22b(+) plasmid and expressed in Escherichia coli BL21 (DE3). The purified recombinant ESAT-6 protein was injected into female BALB/c mice for immunization, and then m-RNA was extracted from the spleen of immunized mice. The anti-ESAT-6 VH/k chain library was assembled by joining of VH and k into the VH/k chain with a 72-bp DNA linker by SOE (splicing by overlap extension) PCR. The scFv library was panned against ESAT-6 using a single round of ribosome display via a rabbit reticulocyte lysate system. Results: ELISA assay showed that one of the selected scFvs had higher affinity against the recombinant ESAT-6 protein. The affinity of the candidate scFv was ̴ 3.74×108 M-1. Conclusion: It could be proposed that the isolated scFv in this study may be useful for the diagnosis of TB. UR - https://ijbms.mums.ac.ir/article_8363.html L1 - https://ijbms.mums.ac.ir/article_8363_16ac0a55e10a7d3e325be59b8f336354.pdf ER -