Open AccessTwo-photon fluorescence correlation microscopy combined with measurements of point spread function; investigations made in human skin
Author(s) -
Stina Guldbrand,
Carl Simonsson,
Mattias Goksör,
Maria Smedh,
Marica B. Ericson
Publication year - 2010
Publication title -
optics express
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.394
H-Index - 271
ISSN - 1094-4087
DOI - 10.1364/oe.18.015289
Subject(s) - fluorescence correlation spectroscopy , human skin , materials science , fluorescence , optics , microscopy , full width at half maximum , point spread function , spectroscopy , two photon excitation microscopy , attenuation coefficient , photon counting , fluorescence microscope , fluorescence spectroscopy , photon , optoelectronics , physics , genetics , quantum mechanics , biology
Two-photon excitation fluorescence correlation spectroscopy (TPFCS) has been applied in connection to measurements of the point spread function (PSF) for quantitative analysis of sulphorhodamine B (SRB) in excised human skin. The PSF was measured using subresolution fluorescent beads embedded in the skin specimen. The PSF, measured as full width at half maximum (FWHM) was found to be 0.41 +/- 0.05 microm in the lateral direction, and 1.2 +/- 0.4 microm in the axial direction. The molecular diffusion of SRB inside the skin ranged between 0.5 and 15.0 x 10(-8) cm(2)/s. The diffusion coefficient is not dependent on depths down to 40 microm. The fluorophores were found to accumulate on the upper layers of the skin. This work is the first TPFCS study in human skin. The results show that TPFCS can be used for quantitative analyses of fluorescent compounds in human skin.