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Updated: Jan 10, 2026
Author Spotlight: Optimizing CFU Determination for Efficient Assessment of TB Vaccine Efficacy and Antigen Presentation Analysis
Published on: July 28, 2023
Optimizing a Subunit Vaccine of Mycobacterium tuberculosis Using In-Silico and In-Vitro Approaches
Zaiqin Ling1, Muhammad Ahsan Naeem2,3
1Department of Tuberculosis, Shandong Public Health Clinical Center, Shandong University, Jinan, 250013, China.
Background:
The present study addresses the development of a novel subunit vaccine (SV) to combat tuberculosis (TB).
Methods:
The research used immunoinformatics to develop a subunit vaccine with 7 MHC-I, 3 MHC-II, and 7 B-cell epitopes joined by AAV, GPGPG, and KK linkers. It involved Mtb protein Rv0577 and PADRE sequence as an adjuvant. TLR2 binding affinity (Kd, nM) was determined through PRODIGY. In-silico evaluations determined allergenicity, antigenicity, and physicochemical properties. The vaccine was presented in an AAVDj/8 system, intracellular expression was verified, and the copy number was identified using qPCR and qRT-PCR.
Results:
The web tools confirmed the stability, non-allergenicity, and high immunogenicity of the vaccine (0.5673 < 0.4). PRODIGY tool depicted good SV-TLR2 binding (ΔG = -8.8 kcal/mol, Kd = 330 nM) with 59 intermolecular contacts, indicating possible TLR2 activation. Indirect immunofluorescence showed the expression of intracellular proteins. Viral titers, determined by 10-fold serial dilution up to 103, showed a detectable titer, and copy numbers (109/mL-1011/mL) proved productive viral replication and significant vaccine effectiveness.
Conclusion:
This comprehensive methodology, from epitope selection to in-vitro testing, establishes a robust foundation for further exploring and advancing this SV.
