Kinetics and comparison of delta-aminolevulinic-acid-induced endogenous protoporphyrin-IX in single cell by steady state and multiphoton fluorescence imaging

Singaravelu Ganesan; Masilamani Elangovan; Ammasi Periasamy

doi:10.1117/12.424563

24 April 2001 Kinetics and comparison of δ-aminolevulinic-acid-induced endogenous protoporphyrin-IX in single cell by steady state and multiphoton fluorescence imaging

Singaravelu Ganesan, Masilamani Elangovan, Ammasi Periasamy

Author Affiliations +

Proceedings Volume 4262, Multiphoton Microscopy in the Biomedical Sciences; (2001) https://doi.org/10.1117/12.424563
Event: BiOS 2001 The International Symposium on Biomedical Optics, 2001, San Jose, CA, United States

Abstract

Photodynamic Therapy has emerged as a new modality in the treatment of various nonmalignant and malignant diseases. It involves the systemic administration of tumor specific photo-sensitizers with the subsequent application of visible light. This combination causes the generation of cytotoxic species, which damage sensitive targets, producing cell injury and tumor destruction. Although, photofrin is the only photosensitizer currently approved for PDT and tumor detection, its concomitant cutaneous photosensitization poses a significant problem. Hence, δ-aminoleuvulinic acid (δ-ALA) a precursor for the endogenous production of Protoporphyrin IX, through heme biosynthesis pathway, has gained significant importance in the Photodynamic Therapy. Though δ-ALA is present naturally in the cells, exogenous δ-ALA helps to synthesis more of PpIX in the tumor cells, as the fast growing tumor cells take up the administered δ-ALA more than the normal cells. Based on these facts, many invasive studies have been reported on the kinetics of δ-ALA at cellular level by chemical extraction of PpIX from the cells. In the present study we have studied the kinetics of δ-ALA induced PpIX fluorescence from Hela cells by perchloric/Methanol extraction method. However, the amount of PpIX synthesized in the cells at different point of incubation time by noninvasive methods has not been reported. Hence we have also used a noninvasive technique of measuring the kinetics δ-ALA induced PPIX fluorescence from Hela, an epithelial cell derived from human cervical cancer by both single photon (steady state) and multi photon excitation. From the studies it is observed that the δ-ALA induced PpIX is more at 2 hours incubation time for 2 mM of δ-ALA concentration. Further, it is observed that with steady state fluorescence imaging method, the excitation light itself cause the Photodynamic damage, due to the prolonged exposure of the cells than in multi photon excitation, leading to the rounding of cells. This may be due to the activation of PpIX in cells by the excitation light source.

Citation Download Citation

Singaravelu Ganesan, Masilamani Elangovan, and Ammasi Periasamy "Kinetics and comparison of δ-aminolevulinic-acid-induced endogenous protoporphyrin-IX in single cell by steady state and multiphoton fluorescence imaging", Proc. SPIE 4262, Multiphoton Microscopy in the Biomedical Sciences, (24 April 2001); https://doi.org/10.1117/12.424563

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