Liu, Hua Qiang
(2024)
Development Of Tcr-like Single
Domain Antibody By Incorporating
Immunoglobulin Isotypes For Latent
Tuberculosis Diagnostics And
Therapeutics Application Against
Heat Shock Protein 16 Kda Peptide
Presented By Hla-a2.
PhD thesis, Universiti Sains Malaysia.
Abstract
Heat shock protein 16 kDa (HSP 16 kDa) is essential for the survival of latent
Mycobacterium tuberculosis. Therefore, a peptide-MHC presentation of HSP 16
kDa could be a potential diagnostic and therapeutic target for latent tuberculosis
(LTB). This study aimed to generate different immunoglobulin isotype pairings on a
TCR-like single-domain antibody (sDAb) and subsequently investigate its
diagnostic and therapeutic potential in LTB, utilizing a model cell presenting the
target peptide. A previously generated TCR-like sDAb that can bind to HSP 16 kDa
was first fused to a human IgG1, IgG2, IgG4, IgA, IgD, IgE, and IgM Fc-receptor
via a linker. The fusion product, sDAb-Fc, was expressed in HEK293-F cells and
subsequently purified. Its diagnostic potential was investigated via cell-based
ELISA utilizing MCF-7 cells peptide-pulsed with HSP 16 kDa peptides.
Antibody-dependent cell-mediated cytotoxicity (ADCC) of MCF-7 cells was also
conducted to investigate its therapeutic potential. Finally, TCR-like sDAb-IgG1,
IgG2, and IgG4 were successfully produced transiently in HEK293 F cells and
purified using protein A chromatography.
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