miniaturized-sample-preparation-for-transmission-electron-microscopy

Miniaturized Sample Preparation for Transmission Electron Microscopy

Due to recent technological progress, cryo-electron microscopy (cryo-EM) is rapidly becoming a standard method for the structural analysis of protein ...

University of Basel

The fight against most infectious diseases, including malaria, is often hampered by the emergence of drug resistance and lack or limited efficacies of vaccines. Therefore, new drugs, vaccines, or other strategies to control these diseases are needed. Here, we present an innovative nanotechnological strategy in which the nanostructure itself represents the active substance with no necessity to release compounds to attain therapeutic effect and which might act in a drug- and vaccine-like dual function. Invasion of Plasmodium falciparum parasites into red blood cells was selected as a biological model for the initial validation of this approach. Stable nanomimics-polymersomes presenting receptors required for parasite attachment to host cells-were designed to efficiently interrupt the life cycle of the parasite by inhibiting invasion. A simple way to build nanomimics without postformation modifications was established. First, a block copolymer of the receptor with a hydrophobic polymer was synthesized and then mixed with a polymersome-forming block copolymer. The resulting nanomimics bound parasite-derived ligands involved in the initial attachment to host cells and they efficiently blocked reinvasion of malaria parasites after their egress from host cells in vitro. They exhibited efficacies of more than 2 orders of magnitude higher than the soluble form of the receptor, which can be explained by multivalent interactions of several receptors on one nanomimic with multiple ligands on the infective parasite. In the future, our strategy might offer interesting treatment options for severe malaria or a way to modulate the immune response. 

Nanomimics of host cell membranes block invasion and expose invasive malaria parasites.

We present a sample preparation method for cryo-electron microscopy (cryo-EM) that requires only 3-20nL of sample to prepare a cryo-EM grid, depending on the protocol used. The sample is applied and spread on the grid by a microcapillary. The procedure does not involve any blotting steps, and real-time monitoring allows the water film thickness to be assessed and decreased to an optimum value prior to vitrification. We demonstrate that the method is suitable for high-resolution cryo-EM and will enable alternative electron microscopy approaches, such as single-cell visual proteomics.

Andrej Bieri

Center for Cellular Imaging and NanoAnalytics (C-CINA),
Biozentrum,
Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum

University of Basel

Miniaturized Sample Preparation for Transmission Electron Microscopy

Andrej Bieri

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Center for Cellular Imaging and NanoAnalytics (C-CINA),Biozentrum,Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum

University of Basel

Miniaturized Sample Preparation for Transmission Electron Microscopy

Center for Cellular Imaging and NanoAnalytics (C-CINA),
Biozentrum,
Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum