Basic fibroblast growth factor (bFGF), also known as FGF-2, is a crucial member of the fibroblast growth factor family, involved in a variety of biological functions including cellular proliferation, wound healing, angiogenesis, intercellular signalling, and cell differentiation, In contemporary stem cell research, serum-free media enriched with various additives and growth factors are employed, and among these, bFGF being particularly significant. Despite its extensive potential applications, the clinical utilization of bFGF is limited due to its instability, especially in aqueous environment. Therefore, a thorough investigation of the protein's structural integrity and stability is essential. This study focuses on the expression, purification, and characterization of bFGF for structural and stability analysis through biophysical methods. Intrinsic fluorescence measurement indicated a structural alteration surrounding the tryptophan residue, while circular dichroism (CD) analysis showed a decrease in the protein’s secondary structure. The differential scanning calorimetry (DSC) used for stability analysis. Furthermore, the study aims to evaluate the biological activity of the protein in cellular context. For this purpose, gold nanoparticles were synthesized. The results from the Cell Migration Assay indicated that the proliferation of HT29 cells was enhanced following treatment with bFGF in conjunction with gold nanoparticles. Also, a MTT assay was conducted.