Polarization effects on fluorescence measurements

Abstract

Po lariza tion effects on fluorescence measurements a re a function of four independent variab les.The fi rst is F, the polarization ratio of th e exciting light which reaches the sample. The second is r, the emission anisotropy of t he sample, which is the po larization " response" of the sample to plane po larized exciting light. The third is C, the polarization ratio of the emission detection system, which is the ratio of the sensitivities of the detection system to vert ica lly and horizontally polarized light. The fourth is ll', the viewing angle, which is the angle bet ween the direction of the propagation of the exciting light and the direction from which the emission is being detect ed.T he intensity and the degree of po larization of the fluorescence emission that the sample exhibits are func tions of F , r, and a , whi le the actual readings obtained with a typica l spectrofluorimeter are functions of al l four variab les, F, r, ll', a nd C. A theoret ica l analysis is made taking all these factors into account, and proper mathematica l models are developed for the different modes of operation in which a fluorimeter can be used. These are verified experimentally with data obtained for a sample which has a high degree of emission anisotropy (Ni le Blue A Perchlorate in glycerol). A recently des igned goniospectrofluorime ter was used. Calibration procedures are deve loped and recommendations are made for modes of operation and fluorescence standards.

Fluorescence behaviour of highly concentrated rhodamine 6G solutions

“…The fluorescence signal is independent of molecular reorientation if the polarizer sheets are oriented under an angle of <j> = arctan (2 1/2 ) = 54.74° (e.g. PI vertical, P2 at angle 0 = 54.74° to the vertical axis) [18,19]. The fluorescence depolarization is obtained by orienting alternately both polarizers parallel (PI and P2 vertical) and perpendicular (PI vertical, P2 horizontal).…”

Intersystem-crossing and excited-state absorption in eosin Y solutions determined by picosecond double pulse transient absorption measurements

“…The fluorescence is collected at right angles to the excitation light propagation direction. A polarizer is inserted in the fluorescence path for magic angle polarization detection [40] (vertically polarized argon ion laser, polarizer in the fluorescence path at an angle of 54.7° to the vertical direction). A monochromator is scanned over the fluorescence wavelength region.…”

Concentration-dependent absorption and emission behaviour of a pyrimidonecarbocyanine dye in hexafluoroisopropanol

“…In case of negligible reorientation of the excited molecules the fluorescence anisotropy is r F = 0.4 for parallel orientation of absorption and emission dipole moment, and r F = -0.2 for perpendicular orientation between absorption and emission dipole moment [8,9]. In case of fast reorientation of the excited molecules within the fluorescence lifetime, r F , the fluorescence anisotropy is r F = 0.…”