Two-photon absorption cross-sections and time-resolved fluorescence imaging using porphyrin photosensitisers

Abstract

Three porphyrin systems have been characterised for use in two-photon fluorescence imaging of biological samples. We have determined the two-photon absorption cross sections (sigma(2)) of the di-cation, free-base and metallated forms of hematoporphyrin derivative (HpD), hematoporphyrin IX (Hp9) and a boronated protoporphyrin (BOPP) using the open-aperture Z-scan and the two-photon induced fluorescence (TPIF) techniques at an excitation wavelength of 800 nm. The insertion of either protons or a metal ion into the macrocycle is shown not to significantly influence the sigma(2) of the porphyrins. Two-photon time-resolved fluorescence images of C6 glioma cells transfected with a free-base form of the BOPP have been obtained as a function of the porphyrin concentration. These studies reveal a maximum useful porphyrin concentration for fluorescence imaging purposes of approximately 30 microg mL(-1).

Fluorescence Lifetime Imaging of Propranolol Uptake in Living Glial C6 Cells

“…For comparison, a second determination of σ 2 for propranolol was made with 2PE at 628 nm using 9-chloroanthracene as a standard with σ 2r = 0.20 GM at 630 nm [25]. Using again the method outlined by Mathai et al [24], a value of σ 2 for propranolol of 0.22 GM at 628 nm was obtained which is in excellent agreement with the value of 0.17 GM units obtained with tryptophan as the standard. Fluorescence lifetimes of propranolol under various conditions were measured using two-photon excitation at 630 nm with the microscope system and placing a drop of solution on a coverslip mounted on the thermostatically controlled microscope stage.…”

“…Under these conditions, propranolol fluorescence intensity exhibits the anticipated quadratic dependence on laser power. two-photon crosssections (σ 2 ) for propranolol as the sample (s) at 590 nm and 630 nm were obtained by the method outlined by Mathai et al (2007) [24] using a reference fluorophore (r) with a known value of σ 2r using (3.1), in which I represents the measured fluorescence intensity in the two-photon experiment, c the concentration, φ the fluorescence quantum yield, and FI λ the fluorescence intensity of the fluorophore in the one-photon excited spectrum at the wavelength λ used in the two-photon experiment relative to the integrated spectral intensity ΣFI:…”

A Metallocorrole with Orthogonal Pyrrole Rings

“…Aromaticity is not only a fascinating electronic-structural phenomenon, responsible for a variety of spectroscopic properties such as intense one-and two-photon absorption, [1,2] aromatics, by virtue of their stability, lend themselves to a host of technological applications, such as photovoltaics, [3] liquid crystals, [4] biological imaging agents, [5] and two-photon absorbers. [2] Nonplanar [6] and chiral [7] aromatics, on account of their topologies, are a particularly intriguing class of molecules, notable not only for their optical/chiroptical properties but also as tools for molecular/chiral recognition.…”

Fluorescence lifetime imaging of E-combretastatin uptake and distribution in live mammalian cells

“…18 Values of σ 2 were measured using a 9-chloroanthracene (9ClA) as a reference fluorophore 18 with an appropriate fluorescence spectrum and measured 2PE cross-sections. 19 The values of σ 2 at 620 nm for E-CA4 and E-CA4F in DCM were determined to be 2.1 ± 0.5 and 2.75 ± 0.6 GM units respectively.…”