Quantitative measurement method for fluorescence lifetime and fluorescence dynamic anisotropic parameters

Abstract

The invention relates to a measuring method for fluorescence characteristics of a fluorescence product and particularly relates to a method for quantitative measurement method for fluorescence lifetime and fluorescence dynamic anisotropic parameters. The method comprises the steps of: exciting a product by a pulse or sine polarization exciting light to produce fluorescence; detecting intensity of polarized fluorescence which is parallel to the pulse or sine polarization exciting light and intensity of polarized fluorescence which is perpendicular to the pulse or sine polarization exciting light respectively with a photoelectric detector; and calculating the average life, the fluorescence anisotropy at the initial time, the fluorescence anisotropy at the infinite time and fluorescence molecule rotational correlation time of the sample. With the adoption of the method provided by the invention, the fluorescence lifetime of all samples can be effectively and quantitatively measured; moreover, the invention provides a quantitative measurement method for fluorescence parameters of the fluorescence product, so the method has great application value for quantitative study of the interaction between large biological molecules, such as proteins, in the living cell.

Description

technical field [0001] The invention relates to a method for measuring fluorescence characteristics of fluorescent samples, in particular to a method for quantitatively measuring fluorescence lifetime and fluorescence dynamic anisotropy parameters. Background technique [0002] Due to the emergence of fluorescent protein molecules (FP), the use of fluorescent labeling and fluorescence measurement techniques to study the biophysical chemistry of living cells and biological tissues has become one of the hotspots in life science research. With the development of modern optics and computer technology, it has been discovered that fluorescence anisotropy and fluorescence lifetime measurement techniques can break through the limitation of Abbe's principle and characterize the interaction between biological macromolecules inside living cells. As described by F.S.Wouters, "The physics and biology of fluorescence microscopy in the life science", in Contemporary Physics (2006), Vol.47,...

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Problems solved by technology

[0007] Jameson.D.M, Gartton, E.and Hall.R, "The measurement and analysis of heterogeneous emissions by multifrequency phase and modulation fluorometry", Appl. Spectrosc. Rev. 20.55-106 (1984) and Joseph R. Lakowicz, Principles of Fluorescence Spectroscopy.(Springer, New York, 2006), 3rd ed proposed a method for quantitatively measuring the fluorescence lifetime of the sample, but for the complex fluorescence process of the sample, these measurement methods cannot effectively quantitatively measure the fluorescence lifetime, while for the fluorescence dynamic anisotropy Anisotropic parameter r 0 , and Quantitative measurement, no specific measurement method has been given yet

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