Justin Taraska, Kem Sochacki and Jenny Hinshaw have long been interested in biological processes like endocytosis that occur at the plasma membrane. Their collaborative team at the US National Institutes of Health wanted to develop a strategy that would overcome existing limitations and improve the study of plasma membrane proteins with cryoET.
“We developed a fast and cost-effective way to prepare ultrathin plasma membranes and used a chemically inducible shape-based protein probe to identify specific proteins within samples,” say the team. To prepare ultrathin plasma membrane sections, the researchers relied on a technique called unroofing. Although they note that unroofing has been used with electron microscopy for ~50 years, their goal “was to push unroofing to the next level — to reach atomic-scale resolution, peer inside and beneath vesicles, and directly observe the proteins that associate with the plasma membrane.”
