adsorption of blood proteins on biomaterial in thrombogen-Total internal reflection fluorescence (TIRF) was applied to esis research (4); the binding to and triggering of living study adsorption of bovine serum albumin (BSA) from a thermocells by hormones, neurotransmitters, and antigens (5); cell dynamically good solvent commonly used as a buffer for BSA adhesion to various surfaces (6); the mechanism of electron crystallization. BSA adsorption is shown to exhibit heteroenergettransport in mitochondrial and photosynthetic membranes ics, which is possibly a general property of protein interfacial be- ; reaction rate enhancement with membrane receptors by havior. For example, BSA adsorption on quartz, SnO 2 , and alkylnonspecific adsorption and surface diffusion of ligands (8); modified surfaces demonstrates broad continuous distributions of immobilized enzymes and antibodies for biosensors and dienergies of adsorption activation and protein-surface binding. For agnostic assays (9); and macromolecule self-assembly at heteroenergetic adsorption, the type of the isotherm and kinetics depends on the type of the energy distribution functions and the interfaces for molecular electronics applications (10). One type of correlation between the energies. In the case of BSA adsorpanalytical technique that has proven to be versatile and capation on quartz and SnO 2 surfaces, protein interfacial behavior is ble of providing a wealth of data on protein interfacial behavconsistent with a model suggesting rectangular distributions both ior is total internal reflection fluorescence (TIRF) (1, 2). of activation energies of adsorption and energies of protein-sur-This method has been successfully applied to studies associface binding. In the case of an alkyl-modified surface, BSA adsorpated with protein adsorption (1-4), surface orientation tion kinetics is interpreted in terms of a heteroenergetic model (11), lateral diffusion (2), rotational mobility, and conforsuggesting initial adsorption at selected adsorption sites with maximational changes of proteins at interfaces 13). Surface mum rate constants. The model also suggests that lateral diffusion selectivity is achieved in TIRF by detecting only the evanesquickly redistributes the molecules from the initial adsorption sites cent wave excited fluorescence which originates within apto the sites with higher binding energies. The combination of TIRF proximately the first 100 nm from the waveguide surface.
spectroscopy with an electrochemical system was used to study the effect of surface charge on protein adsorption on a transparent TIRF allows in situ, real-time, nondestructive, and highly SnO 2 electrode. BSA behavior on the SnO 2 electrode was found to sensitive investigation of surface behavior even in the prescorrelate with surface hydrophobicity and indifferent to charges ence of high concentrations of protein in the bulk of solution.
both of the surface and the protein. Hydrophobic interactions seem
We report here a new TIRF application which may be to be the principal driving force in determining the behavior of useful for investigating molecular mechanisms of protein BSA at the solid-liquid interface and likely plays an important crystallization. In protein crystallogenesis, attachment of disrole in protein crystallogenesis.