My project focuses on the synthesis and intracellular detection of transition metal complexes, specially metal carbonyl complexes of the M(CO)3 type. We recently showed that such compounds could be mapped in cellulo using their infrared signature.
One example of the mappings obtained by infrared spectromicroscopy on the SMIS beamline is shown in Figure 1. MDA-MB-231 cells (breast cancer) were incubated with P89, a metal carbonyl analogue of tamoxifen (molecule developed in the Friedel laboratory, ENSCP). The maps were generated by measuring and plotting the integral of specific infrared bands (hot spots were obtained). P89 was thus detected by one of its absorption bands at 1925 cm-1 (image b). The nucleus presents different infrared signatures, including the absorption of amides (amide I band at 1650 cm-1) reflecting a high concentration of DNA folding proteins, histones (image c). When both maps were superimposed (image d, overlay in yellow) we were able to deduce the localization of P89 in the nucleus. A correlative study with fluorescence microscopy (using DAPI to label the nucleus) backed this observation.
Figure 1. Two MDA-MB-231 breast cancer cells (cells 1 and 2) incubated with a metal-carbonyl tamoxifen analog P89 (10 µM, 1 h, 37 °C, 5% CO2). (a) Bright field image (scale bar 10 µm). IR mappings recorded at the SMIS beamline: (b) P89-band hot spot (1925 cm—1, green); (c) amide I-band hot spot (1650 cm—1, red); (d) P89-band hot spot (green), amide I-band hot spot (red), overlay (yellow). Pixel size: 6 x 6 µm2.
Given the diversity of problems encountered in biology, it may be useful to have "multimodal" sensors, detectable in complementary ways. There are examples in the literature of molecules combining fluorescence and MRI or fluorescence and radio emissions. The majority of these objects are of high molecular weight and each modality is assured by a different fragment. We are looking to synthesize probes the multimodality of which is based on a single molecular core (we suggest the acronym SCoMPI for "Single Core Multimodal Probe for Imaging") to cause the least disturbance to the biological system under study.
At the moment we are interested in bimodal probes, both infrared active and luminescent. Fluorescence microscopy is a technique widely adopted by biologists and a wide range of organelle markers is available. Infrared spectroscopy, in turn, involves radiation that is much less damaging to cells and tissues and allows the use of endogenous signals to detect certain organelles (such as the absorption of phosphates and amides to identify the nucleus).
The poster presents the physicochemical characteristics of one of these probes and its potential as a bimodal marker for subcellular imaging.
Its infrared spectral signatures and fluorescence emissions are outside the intrinsic responses of the cell, allowing an unambiguous detection of the compound in cellulo.
From an imaging point of view, a first objective was to show that the information provided by the two spectroscopies was coherent, that the infrared and luminescence maps could be superimposed, giving the same cellular distribution for the probe. We provided evidence of perinuclear localization using both methods, showing the relevance of SCoMPI for bioimaging. Colocalization studies with fluorescence markers showed that the probe was localized to the Golgi apparatus.
The results, for which the poster provides a summary, have just been published in the journal Chemical Communications
Clède S, et al, A Rhenium tris-Carbonyl Derivative as a Single Core Multimodal Probe for Imaging (SCoMPI) Combining Infrared and Luminescent Properties, Chem. Commun. 2012, 48,7729-7731