My watch list


Membrane nano-tomography in living cells

Label-free evanescent microscopy enables full-field and real-time tracking of membrane processes without signal fading and cell perturbation.

08-Dec-2014: Membranes play a pivotal role in numerous cell mechanisms, in particular for internalization, adhesion and motility studies. In terms of optical imaging of the membrane, special configurations are needed to remove the light coming from the inner part of the cell. French scientists now show that through-the-objective evanescent microscopy (epi-EM) is a powerful technique to image membranes in living cells.

In label-free evanescent microscopy (EM), configurations similar to total internal reflection fluorescence (TIRF) have been proposed: prism-based or through-the-objective. However, in the latter case, these evanescent techniques have not spread much, with a relative preference for the prism-based configuration also called total internal reflection microscopy (TIRM). The team led by Pierre Bon chose the through-the-objective based configuration (epi-EM), which enabled super-axially resolved tomographic reconstruction of the basal membrane of label-free living cells. The implementation of epi-EM only required an easy to settle illumination/collection scheme on a standard inverted microscope. Only a high-NA objective (NAobj > 1.33) was needed for living biological sample studies and a spatial filter on the epi-illumination arm in order to reject under-critical angle illumination.

Either bead calibration or a multilayer Fresnel model could be used to retrieve nanometric position. Based on a multilayer Fresnel model, the team was able to retrieve the membrane/interface distance with 10 nm precision. The researchers applied this nano-axial tomography to retrieve quantitative information on invagination dynamics of living cell membranes. They studied the membrane elevation map of living cells (Wt HEK-293) during 15 minutes at one frame per second without perturbing the sample.

The results demonstrate that epi-EM gives easily access to axially super-resolved images of unlabeled microscopic samples with almost no microscope modification, and at least a doubled lateral resolution compared to classical TIRM. A study can be of any duration as the signal level is not sensitive to any fluorophore stability dependence and the photoxicity is very low as barely any light is absorbed by the sample. The scientists are convinced that this technique will be useful for cell motility and adhesion studies when the sample cannot be modified (ex. stem cells) or when very fast and/or long studies are required.

Original publication:
P. Bon, T. Barroca, S. Lévèque-Fort, E. Fort; Label-free evanescent microscopy for membrane nano-tomography in living cells, J. Biophotonics 7:11-12, 857-862 (2014); doi: http://dx.doi.org/10.1002/jbio.201300095

Facts, background information, dossiers

  • tomography

More about Wiley-VCH

  • News

    For a Better Contrast

    Magnetic resonance imaging (MRI) has emerged as one of the most powerful clinical imaging tools because of its superb spatial resolution and soft tissue contrast, especially when using contrast agents. In the European Journal of Inorganic Chemistry, scientists have presented a new kind of n ... more

    Garlic ingredient from the lab bench

    Fresh garlic extracts contain a variety of healthy organosulfur compounds, among which ajoene forms a major oil-extractable ingredient. Now, chemists in the United Kingdom have synthesized ajoene from readily available components for the first time. The results show that ajoene is accessibl ... more

    Smart Fluorescent Dyes

    Controlling the excited electronic states in luminescent systems remains a challenge in the development of fluorescent and phosphorescent dyes. Now, scientists in Japan have developed a unique organic fluorophore that changes its emission color without loss of efficiency when externally sti ... more

q&more – the networking platform for quality excellence in lab and process

The q&more concept is to increase the visibility of recent research and innovative solutions, and support the exchange of knowledge. In the broad spectrum of subjects covered, the focus is on achieving maximum quality in highly innovative sectors. As a modern knowledge platform, q&more offers market participants one-of-a-kind networking opportunities. Cutting-edge research is presented by authors of international repute. Attractively presented in a high-quality context, and published in German and English, the original articles introduce new concepts and highlight unconventional solution strategies.

> more about q&more

q&more is supported by:


Your browser is not current. Microsoft Internet Explorer 6.0 does not support some functions on Chemie.DE