Probing Relative Humidity Impact on Biological Protein Bovine Serum Albumin and Bovine Submaxillary Gland Mucin by Using Contact Resonance Atomic Force Microscopy

[Image: see text] In biomaterials, a substantial amount of research has been placed on the mechanical properties of biomolecules and their interactions with body fluids. Bovine serum albumin (BSA) is a widely studied model protein, while bovine submaxillary gland mucin (BSM) is another cow-derived protein frequently employed in research. Films were examined with contact resonance atomic force microscopy (CR-AFM), and the results showed that the mechanical characteristics of the films were affected by the relative humidity. We quantitatively analyze the viscoelasticity of these proteins after they have been subjected to humidity by measuring the resonance frequency and quality factor. The findings indicate that prolonged humidity exposure has a different effect on the mechanical properties of BSA and BSM films. The results show that after exposure to humidity, the resonance peaks of BSA shift to the left, indicating stiffness, while those of BSM shift to the right, indicating hydration. Moreover, BSM’s hydration is caused by relative humidity, leading to a constant increase in resonance frequency and material softness. Contrarily, BSA showed a decrease in contact resonance frequency due to ongoing strain-induced deformation, indicating increased material stiffness. The findings have significance for the design and development of biomaterials for a variety of applications, such as the delivery of drugs, the engineering of tissue, and the development of biosensors. Our research demonstrates that CR-AFM has the potential to become a non-invasive and sensitive method that can be used to characterize the mechanical characteristics of biomolecules and their interactions with bodily fluids.