TOTAL INTERNAL REFLECTION FLUORESCENCE (TIRF) AS A QUANTITATIVE PROBE OF PROTEIN ADSORPTION.

Total internal reflection fluorescence (TIRF) is a powerful new method for probing protein adsorption at solid-liquid interfaces. A quantitation method for determining the amount of adsorbed protein on hydrophilic glass and quartz surfaces has been developed. This method utilizes fluorescent, non-adsorbing dextran as a calibration molecule which approximates the diffusive properties of the protein under study. Through the use of conventional bulk solution fluorescence of both protein and calibration molecules in conjunction with a TIRF calibration, graphical quantitation of the surface concentration of adsorbed protein can be performed. Quantitative results can be obtained either via a graphical procedure using fluorescence calibration standards or by a numerical method which accounts for the characteristics of the decaying evanescent, interfacial wave.

Duke Scholars

Author William M. Reichert Biomedical Engineering