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Internalization of Luminescent Calcium Phosphate Nanoparticles By Human Cancerous Pancreatic Ductal Cells (Capan-1 Line)

Posted on: Thursday, 10 June 2004, 06:00 CDT

VT-2021

Fluorescent organic molecules such as fluoresceine are widely used as biological probes. However, the photobleaching of such molecules occurs quickly. Therefore, tests have been carried out with fluorescent semiconductor nanocryslals (CdSe-CdS, CdSe-ZnS core- shell nanoparticles) on which proteins have been grafted. Their possible use as biological labels was shown. The particles must be nanometric sized so as to facilitate their internalization by the cells. In order to have a more biocompatible material, we have synthesised luminescent probes with calcium phosphate doped with a rare earth, europium. Calcium phosphates of apatitic structure are similar to the mineral part of calcified tissues in living organisms (bones, teeth): their structure and composition are very close, and they are biocompatible and biodegradable. Europium leads to a maximum emission peak at 1 = 614 nm and is much more photostable than common organic molecules used for cell characterisation. Europium ions (2% atomic Eu/(Eu+Ca)) have been introduced in apatitic tricalcium phosphate during synthesis in partially aqueous medium at body temperature (37 C). The luminescence excitation spectrum of these materials shows that they can be excited with a visible wavelength, which is a more advisable condition to use them in biological applications. To demonstrate the internalization of the europium-doped nanoparticles by a cell system, human cancerous pancreatic ductal cells (Capan-1 line) were maintained in the presence of doped calcium phosphate powder (30 min, 37C). Analysis by confocal laser scanning microscopy (excitation[lambda] = 488 nm) showed the presence of fluorescent structures scattered in the cytoplasm of treated cells. By transmission electron microscopy, electron-dense crystallites were observed on the surface of plasma membranes and in the matrix of cytoplasmic vesicles (around 100 nm to 2 m diameter) in treated Capan-1 cells. Electron microdiffraction analysis confirm that these crystallites still had their original apatitic structure. Taken together these results indicated that europium-doped nanoparticles could be internalized in a cell system and readily visualized by microscopy. These nanoparticles thus constitute interesting luminescent probes for the targeting of various molecules to specific cells.

M. FANJUL1,2, A. Doat3, F. Pelle4, E. Hollande1,2, and A. Lebugle3. 1 Laboratoire de Biologie Cellulaire et Moleculaire des Epitheliums, Universite Paul Sabatier, 38, rue des 36 Ponts, 31400 Toulouse, France; 2 Unite INSERM U531, Biologie et Pathologie Digestive, CHU Rangueil Bat L3, 31403 Toulouse, France; 3 CIRIMAT, UMR 5085, Physico-Chimie des Phosphates, ENSIACET-INPT, 31077 Toulouse, France; and 4Groupe d'Optique des Terres Rares, UMR 7574 Materiaux Inorganiques, I. place A. Briand, 92195 Meudon Cedex- France. E-mail: fanjul@lmtg. ups-tlse.fr

Copyright Society for In Vitro Biology Spring 2004

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