Related Experiment Video
Updated: Aug 24, 2026
Separation and Fractionation of Culture Filtrate Proteins (CFPs) from Mycobacterium tuberculosis
Published on: July 11, 2025
[Molecular cloning, purification, and serological characterization of four specific antigens of Mycobacterium
Lei Xu1, Hui-yun Li, Li-heng Ma
1Department of Biochemistry, Medical College, Tongji University, Shanghai 200092, China.
Objective:
To express the 14,000, 37,000, and 6,000 early secretory antigenic target (ESAT-6) and mtb81 antigen genes in bacteria, and to purify the product and determine their activity.
Methods:
The 14,000, 37,000 , ESAT-6, and mtb81 antigen genes were amplified from Mycobacterium tuberculosis genomic DNA by polymerase chain reactions and cloned into pGEX 4T-1 expression vector. BL21 strain of Escherichia coli was transformed with the recombinant vectors and induced to express recombinant proteins. The proteins were purified by affinity chromatography. The biological activity of purified proteins were estimated by enzyme-linked immunoabsorbant assay (ELISA).
Results:
The BL21 strains of Escherichia coli with recombinant vectors showed high level of 14,000, 37,000, ESAT-6, and mtb81 gene expressions after induction. The products were purified successfully and showed high antigenicity and specificity. The sensitivity of 38,000, 14,000, ESAT-6, and mtb81 were 54%, 60%, 44%, and 36%, respectively.
Conclusion:
The expressions and purifications of recombinant 14,000, 37,000, ESAT-6, and mtb81 antigens with natural activity facilitate their research and application.
