Adsorption of bovine serum albumin on fused silica: Elucidation of protein-protein interactions by single-molecule fluorescence microscopy.
The adsorption of bovine serum albumin (BSA) on fused silica at pH 4.7 was studied at the single molecules level by total-internal-reflection fluorescence microscopy. This pH value was the isoelectric point of BSA. At low [BSA] of 20pM, protein molecules adsorbed as monomers. At intermediate [BSA] of 500pM, protein molecules adsorbed as clusters of about five monomers on average. Both monomers and clusters had adsorption rate coefficients of the order 10(-7)ms(-1) and desorption rate coefficients of about 2x10(-2)s(-1). The respective steady-state coverage was about 10x higher than that at neutral pH, presumably because of the more favorable BSA-silica electrostatics. At pH 4.7 and with [BSA] higher than 100nM, adsorption begot further adsorption to produce nonlinear isotherms. The coverage at 1microM BSA was 2.5x that of the linearly extrapolated coverage. This suggests that at pH 4.7, solute-adsorbate affinity was the dominant factor that explains the enhanced adsorption observed in ensemble measurements.
Yeung
K M
KM
Department of Physics, Hong Kong Baptist University, Kowloon Tong, China.
Lu
Z J
ZJ
Cheung
N H
NH
D000327:Adsorption; D055592:Biophysical Phenomena; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D008856:Microscopy, Fluorescence; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D012822:Silicon Dioxide / Q000737:chemistry*; D055672:Static Electricity; D013499:Surface Properties
D012710:Serum Albumin, Bovine; D012822:Silicon Dioxide