Two-photon characterization and microscopy of porphyrin photosensitisers
2007
The benefits of two-photon fluorescence microscopy of biological samples are vast arising from the utilisation of low
energy light. The two-photon absorption cross sections (σ 2 ) of the di-cation free-base and metallated forms of
hematoporphyrin derivative (HpD), hematoporphyrin IX (Hp9) and a boronated protoporphyrin (BOPP) are obtained to
ascertain their effectiveness as fluorophores for use in two-photon microscopy. The open-aperture Z-scan and the two-photon
induced fluorescence (TPIF) techniques, each capable of providing information regarding the nonlinear
absorption, are employed to determine σ 2 of the various porphyrins at an excitation wavelength of 800 nm.
A significant disparity in the determined values of σ 2 using the two methods is observed. This is largely attributed to the
common requirement of higher concentrations used in the open aperture Z-scan method compared with TPIF techniques.
Values of σ 2 obtained from the Z-scan experiments are in the order of 10 GM, whilst those obtained from the TPIF
experiments are in the order of 200 GM. Insertion of either protons or metal ions into the macrocycle does not enhance
the σ 2 of the porphyrins. Successful two-photon induced fluorescence imaging of BOPP free-base loaded G6 glioma cells
is achieved, confirming the usefulness of this porphyrin in two-photon microscopy.
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