Total Internal Reflection Fluorescence Microscopy

This lecture covers the principles and applications of Total Internal Reflection Fluorescence Microscopy (TIRFM). It explains how TIRFM uses evanescent waves to selectively excite fluorophores near the coverslip surface, providing high-contrast images. The lecture discusses the limitations of conventional fluorescence microscopy and the conditions required for Total Internal Reflection (TIR). It also explores the concept of evanescent waves in TIR, the penetration depth of the evanescent wave, and various fluorescence microscopy techniques. Furthermore, it delves into the details of Fluorescence Recovery After Photobleaching (FRAP) method, its applications, kinetic analysis, and equipment. The lecture concludes with examples of FRAP experiments and the measurement of diffusion coefficients using FRAP.