6th ESACP Congress, Heidelberg, April 7-11, 1999

A014
RAMAN SPECTROSCOPY OF VIABLE CELLS AND TISSUES
Manfait M, Sockalingum GD, Morjani H, Beljebbar A

Unité MEDIAN, IFR53, UFR de Pharmacie, Université de Reims Champagne-Ardenne, France

Raman spectroscopy is recognised as one of the most effective approaches in pathology diagnosis. This technique has been used to differentiate between sensitive and MDR-resistant cancer cells. Several cell lines including K562, CEM, LR73 and MCF7 (MDR phenotype) and HL60 and J82 cells (non-MDR phenotype) have been investigated. Spectral analysis of the sensitive and resistant cell lines has allowed to identify the differences proper to the MDR and non-MDR-phenotype. In the case of MDR phenotype, sensitive cells exhibit a higher intensity than the resistant counterpart, in particular in the amide I and III regions. This was not the case with non-MDR phenotype. Also, cells exhibiting MDR-phenotype obtained both by drug selection and transfection with mdr gene, are included. Data show that it can be possible to identify the spectral characteristics associated with this resistance mechanism (profile, percentage of proteins and lipids). Resolution enhancement methods (deconvolution, derivatives, and curve fitting) have also been used to evaluate the conformational changes in the secondary structure of cell constituents that accompany the MDR phenotype. Surface-enhanced Raman spectroscopy (SERS) has become a powerful and non-invasive probe for investigating the molecular and cellular interaction of drugs with their targets (DNA, receptors, and enzymes). The comparison in-vitro models allow to elucidate the biological effect of the drugs. The development of new types of SERS-active substrates has extended the applicability of this technique to medical diagnosis. Two kinds of SERS active substrates, characterised as "bio-compatible" systems, can be used for investigation on single living cells: colloid suspensions and microelectrodes (point probes), and island films (surface probes). This technique has been used as a rapid, effective and reproducible method to investigate in-situ different cellular compartments for a better localization of the drug inside the cell (free or bound to its pharmacological target).