Current in vitro optical studies of microtubule dynamics tend to rely on fluorescent labeling of tubulin, with tracking accuracy thereby limited by the quantum yield of fluorophores and by photobleaching. Here, we demonstrate label-free tracking of microtubules with nanometer precision at kilohertz frame rates using interferometric scattering microscopy (iSCAT). With microtubules tethered to a glass substrate using low-density kinesin, we readily detect sequential 8 nm steps in the microtubule center of mass, characteristic of a single kinesin molecule moving a microtubule. iSCAT also permits dynamic changes in filament length to be measured with <5 nm precision. Using the arbitrarily long observation time enabled by label-free iSCAT imaging, we demonstrate continuous monitoring of microtubule disassembly over a 30 min period. The ability of iSCAT to track microtubules with nm precision together with its potential for label-free single protein detection and simultaneous single molecule fluorescence imaging represent a unique platform for novel approaches to studying microtubule dynamics.

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Class I MHC molecules have been thought to occur in vivo both as class I MHC heavy chain-beta 2-m heterodimers, which are or are not associated with antigenic peptide, and as free class I MHC heavy chains. Class I MHC molecules are now found also to occur in another type of structure: a heavy chain-heavy chain dimer. Biochemical studies show that heavy chain dimers are disulfide-linked via a conserved cytoplasmic domain cysteine. H-2Ld, H-2Db, and H-2Dd class I dimers fail to react with certain alpha 1 and alpha 2 domain-specific antibodies. Furthermore, although beta 2-m-specific antibodies coprecipitate class I MHC heavy chains, they do not coprecipitate class I MHC heavy chain dimers. Pulse-chase studies show that heavy chain dimer formation occurs at different points in the biosynthesis of class I MHC molecules in beta 2-m+ and beta 2-m- cells: in beta 2-m+ cells, heavy chain dimers form after the class I molecules have traversed the medial Golgi cisternae, whereas in beta 2-m- cells they form immediately. Culturing of beta 2-m+ cells with exogenous beta 2-m prevents the formation of H-2Ld/Db heavy chain dimers. We conclude that dimer formation occurs as a consequence of loss or unavailability of beta 2-m. Class I MHC heavy chain dimerization may provide a mechanism for removal of immunologically dysfunctional molecules.

In testing biological fluids for specific antigens by the method of crossed immunoelectrophoresis in agar gel (Bussard, 1959;Culliford, 1964;Kohn, 1967) and tapered troughs are cut either with a fine scalpel or with a suitable gel punch. For a 1-mm layer of agar, the troughs shown in Fig. 1

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