Single-particle visualization of assembly: I. Dimerization in a planar zone

Single-particle fluorescence microscopy of association/dissociation is required for analysis of biological assembly reactions. Toward achieving this goal, Wang et al. (J. Microsc., 2004, 213, 101-109) used molten agarose to concentrate thermally diffusing particles in a thin zone of solution next to the surface of a coverglass (plane of concentration). The present study details the first real-time, single-particle analysis of the association/dissociation of thermally diffusing particles in the plane of concentration. The test particles were procapsids of bacteriophage λ (radius = 31 nm). Quantification of thermal motion was developed and used to determine whether co-diffusing particles were bound to each other. The data are explained by (1) the presence of a molten agarose-generated barrier that is 93-155 nm from the coverglass surface, and (2) nonrandom orientation of procapsid dimers in the plane of concentration.