Fluorescence Correlation Spectroscopy英文文献.pdfVIP

6 Fluorescence Correlation Spectroscopy Nancy L. Thompson 6.1. Introduction Fluorescence correlation spectroscopy (FCS) is a technique in which temporal fluctuations in the fluorescence measured from a sample of fluores- cent molecules are analyzed to obtain information about the processes that give rise to the fluorescence fluctuations. Since the initial development,(1–3) FCS has been used to measure translational diffusion coefficients,(4–18) chemi- cal kinetic rate constants,(1, 3, 8, 19–21) rotational diffusion coefficients,(22–24) flow rates,(25) molecular weights,(17, 26) and molecular aggregation.(16, 27–29, 30) Measurement of these physical parameters has been used for a range of purposes, including an increased understanding of protein, nucleic acid, and lipid diffusion,(5–7, 14, 15, 18, 23) solution and interfacial kinetics,(20, 21, 31) cell-surface receptor(27, 30) and macromolecular(16, 28, 29) clustering, and muscle contraction(8, 22, 32); the detection of antibodies(10, 12, 31) and viruses(33, 34); and the characterization of focused laser beams.(11, 35) In this chapter the conceptual basis and theory of FCS, aspects of FCS apparatus and experi- mental methods, and most of the previous experimental applications of FCS are described. FCS has been the subject of a number of previous review articles and book chapters.(13, 32, 36–51) 6.2. Conceptual Basis and Theoretical Background The conceptual basis of FCS is illustrated in Figure 6.1. At equilibrium, fluorescent molecules move through a small open regi