Adhesion and stiffness of biotin-superavidin bonds

A non-invasive vibrational spectroscopy technique is introduced and utilized to characterize the average spring constant of a single Superavidin (SAv)-Biotin (Bi).polyethylene glycol (PEG) ligand receptor complex as well as the effective Young's modulus and adhesion of a layer formed by the SAv-Bi.PEG ligand-receptors. In the reported experiments, SAv coated Polystyrene (PS) particles are deposited on a layer of Bi.PEG receptors, bound to a silicon (Si) substrate by silanization. The substrate and the bonded particles are subjected to a pulsed ultrasonic excitation field and their nanometer scale out-of-plane dynamic responses are acquired using a laser vibrometer. The acquired waveforms are processed to obtain the resonance frequencies of the particle motion. Employing a particle adhesion model, the average spring constant of the single ligand-receptor complex and the effective Young's modulus and work-of-adhesion of the SAv-Bi.PEG ligand-receptor layer are extracted from the resonance frequencies. The average spring constant of an individual SAv-Bi.PEG bond is approximated as 0.1-0.4 mN/m. The work-of-adhesion and effective Young's modulus of the SAv-Bi.PEG layer are determined to be 0.54-2.62 J/m² and 0.15-2.80 MPa, respectively. The compressive Young's modulus of the SAv-Bi.PEG layer is estimated as 31.0-58.0 MPa. The current approach provides a direct non-contact measurement technique for the stiffness of single ligand receptor complexes and the adhesion of their interfaces. SAv-Bi bonds and PEG polymers are among the most widely utilized complexes in the pharmaceutical and biological applications. Understanding the mechanical properties of PEG and SAv-Bi is an important step towards optimization of their utilization in practical applications such as biosensors and targeted drug delivery.