13 Nov - A new toy for our EM facility arrived: a new “plunge-freezer”; the Leica EM GP2!!!
A plunge-freezer is an instrument to vitrify (= freeze without ice crystal formation) very thin films of mostly suspensions of isolated protein fractions, vesicles, bacteria or cells on EM-support grids. This process stabilizes the ultrastructure of the sample without the need for chemical fixation. The vitrified grids are observed and imaged at -180°C under low electron dose conditions in a transmission electron microscope.
In general, the plunge-freezing process implies the application of a tiny drop of a suspension by a micropipette onto a grid held by a forceps in the plunge-freezer, incubation of the suspension for a few seconds, blotting off the access of suspension and finally plunging the grid into a liquid nitrogen cooled puddle of ethane close to its freezing point.
The Leica EM GP2 replaces our current - mostly manually operated - model, that has served us well, because the new instrument has a number of important advantages. It offers the possibility to parameterize many factors, such as the relative humidity, incubation time, blotting time and number of blots, and several others. As blotting, one of the most critical steps of the process, can be fully automatic, it is expected that this will highly improve reproducibility. In addition, the excellent control of environmental conditions during the process will reduce contamination of the grids by condensing water vapor greatly.
In summary, we are confident that the acquisition of the Leica EM GP2 plunge-freezer will improve the success rate of the freezing process importantly.
4 Oct - NOBEL prize 2017 for cryo-electron microscopy
The Nobel prize for Chemistry 2017 went to 3 investigators (Jacques Dubochet, Joachim Frank and Richard Henderson) whose scientific contributions have led to our current ability to determine the 3D-structure of biomolecules by cryo-electron microscopy.
Conventional preparation techniques for electron microscopy use chemical preparation of biological samples, inevitably influencing the observed ultrastructure to a more or lesser extent. Cryo-EM at the other hand, uses ultra-rapid cooling of a very thin film of a suspension containing the molecules of interest, vitrifying the sample without formation of ice-crystals that could damage the ultrastructure. By applying mathematical algorithms (single particle analysis) to thousands of images of the molecules of interest in many different orientations in the vitrified suspension, the 3D-structure of the molecules is revealed. By recent technological advances in instrumentation it is now possible to obtain 3D-structures at atomic resolution. This information can contribute to solving many questions in biomedical context. See also www.Nobelprize.org.
Whereas we in Leuven use cryo-EM for elucidating ultrastructural features of lipid structures in particular (exosomes, liposomes, etc.) with less demanding resolution requirements, our VIB-colleagues in Brussels (Rouslan Efremov, Structural Biology, VIB@VUB) will install a high-end cryo-electron microscope (expected 2018) for pursuing 3D-structure of biomolecules as described above.