Single particle reconstruction

Single particle reconstruction is a technique in which large numbers (10,000 - 1,000,000) images of ostensibly identical individual molecules or macromolecular assemblies are combined to produce a 3 dimensional reconstruction. This is a complementary technique to crystallography of biological molecules. As molecules/assembies become larger, it becomes more difficult to prepare high resolution crystals. In single particle reconstruction, the molecules/assemblies in solution are prepared in a thin layer of vitreous (glassy) ice, then imaged on an electron cryomicroscope (see Transmission electron microscopy). Images of individual molecules/assemblies are then selected from the micrograph and then a complex series of algorithms often requiring thousands of CPU-hours is used to produce a full volumetric reconstruction of the molecule/assembly. In the 1990's this technique was limited to ~2 nm resolution, providing only gross features of the objects being studied. However, recent improvements in both microscope technology as well as available computational capabilities now make 0.5 nm resolution possible. The resolution is expected to approach the 0.2 - 0.3 nm resolutions achievable by crystallography over the next decade.