Distribution of ALA metabolic products in esophageal carcinoma cells using spectrally resolved confocal laser microscopy

Jozef Smolka; Anton Mateasik

doi:10.1117/12.697302

21 July 2006 Distribution of ALA metabolic products in esophageal carcinoma cells using spectrally resolved confocal laser microscopy

Jozef Smolka, Anton Mateasik

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Proceedings Volume 6163, Saratov Fall Meeting 2005: Optical Technologies in Biophysics and Medicine VII; 61630Z (2006) https://doi.org/10.1117/12.697302
Event: Saratov Fall Meeting 2005, 2005, Saratov, Russian Federation

Abstract

Aminolevulinic acid (ALA) is an efficient substance used in photodynamic therapy (PDT). It is a precursor of light-sensitive products that can selectively accumulate in malignant cells following the altered activity of the heme biosynthetic pathway enzymes in such cells. These products are synthesized in mitochondria and distributed to various cellular structures [1]. The localization of ALA products in subcellular structures depends on their chemical characteristics as well as on the properties of the intracellular environment [2]. Characterization of such properties is possible by means of fluorescent probes like JC-1 and carboxy SNARF-1. However, the emission spectra of these probes are overlapped with spectral pattern of typical ALA product -protoporphyrin IX (PpIX). Spectral overlap of fluorescence signals prevents to clearly separate a distribution of probes from PpIX distribution what can completely mess the applicability of these probes in characterization of cell properties. The spectrally resolved confocal laser microscopy can be used to overcome this problem. In this study, a distribution of ALA metabolic products in relation to the mitochondrial membrane potential and intracellular pH was examined. Human cell lines (KYSE-450, KYSE-70) from esophageal squamous cell carcinoma were used. Cells were incubated with 1mM solution of ALA for four hours. Two fluorescent probes, carboxy SNARF-1 and JC-1 , were used to monitor intracellular pH levels and to determine membrane potential changes, respectively. The samples were scanned by spectrally resolved laser scanning microscope. Spectral linear unmixing method was used to discriminate and separate regions of accumulation of ALA metabolic products of JC-1 and carboxy SNARF-1.

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Jozef Smolka and Anton Mateasik "Distribution of ALA metabolic products in esophageal carcinoma cells using spectrally resolved confocal laser microscopy", Proc. SPIE 6163, Saratov Fall Meeting 2005: Optical Technologies in Biophysics and Medicine VII, 61630Z (21 July 2006); https://doi.org/10.1117/12.697302

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KEYWORDS

Luminescence

Confocal microscopy

Photodynamic therapy

Linear filtering

Microscopy

Microscopes

Image processing

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