Electrophoretic Studies on Elementary Bodies of Vaccinia

Abstract

It has been found (1) that suspensions of elementary bodies of vaccinia studied by electrophoresis in the Tiselins apparatus yield moving boundaries which differ from those usually observed. Unlike solutions of ordinary proteins or of two of the plant virus proteins (tobacco mosaic (2) and bushy stunt (3)) which on electrolysis display boundaries that remain quite sharp and fiat, suspensions of the elementary bodies of vaccinia show distortions in the form of a stream of the particles moving ahead from the center of the boundaries in the direction of the electric current. Thus, one observes a stream of the suspended material rising in the center of one leg of the cell and a corresponding stream of the solvent descending in the center of the other leg. Reversal of the current results in the reversal of these effects. Because of the asymmetric nature of this boundary disturbance, simple convection arising from the electrical heat generated cannot account for the phenomenon. McFarlane (1) attributed the effect to endosmosis.It will be shown by data presented in this paper that the type of boundaries observed with vaccine virus can be obtained also with particles of a similar size but of an entirely different nature. Furthermore, the mobility of such boundaries in a given medium depends principally on the nature of the surface of the particles, a fact which has long been known to workers using the microscopic electrophoretic method in which the motion of a single particle is studied (4). In addition, the boundary distortions obtained with suspensions of particles will be shown to be artifacts in that they do not result from any characteristic property of the particles themselves but rather from the absence of density gradients of sufficient magnitude at the boundary. Materials and MethodsElementary Bodies of Vaccinia.--Elementary bodies of vaccinia were prepared according to the method of Craigie (5) which has recently been redescribed at length (6). 40 ce. of a stock suspension of virus obtained from one rabbit contained about 8 rag. of dry material (7); thus, the concentration was approximately 0.2 mg. per cc. or 0.2 per

Electrophoretic Studies on Pr8 Influenza Virus

“…Because of the strong opalescence of solutions of influenza virus, it was necessary to employ infrared light and infrared sensitive photographic plates as recommended by Treffers and Moore (17). A marked tendency towards streaming of the protein during electrophoresis, similar to that encountered with vaccinia virus (18) and with derivatives of tobacco mosaic virus (19), was diminished by allowing the boundaries to move at a very slow rate of 0.3 era. per hour.…”

Observations on Mixtures of Elementary Bodies of Vaccinia and Coated Collodion Particles by Means of Ultracentrifugation and Electrophoresis

“…Previous experiments have shown that elementary bodies of vaccinia migrate in the Tiselius apparatus at a rate of about 10 X 10 -6 cm./sec. per volt/cm, in monovalent buffer solution, 0.01 ~t pH 7.9 (2,5), and that 523 collodion particles coated with the purified heat-stable soluble antigen (S) of vaccinia move at approximately the same rate under similar conditions (2). It has also been demonstrated that collodion particles coated with one of the components of normal rabbit serum move at a different rate under these experimental conditions.…”

“…The various materials used and their method of preparation have been described in the preceding publication (2). Mixed suspensions of coated collodion particles and elementary bodies of vaccinia were studied in the Tiselius apparatus by the method previously described (2) and in the analytical ultracentrifuge of Bauer and Pickels (3) by the technique described several years ago (4).…”

“…The collodion particles coated with a component of normal serum moved at a rate of 13 X 10 -5 cm./sec, per volt/cm., while the elementary bodies traveled at their usual speed of 526 10 × 10 -5 cm./sec, per volt/cm. Differences in staining characteristics of coated collodion particles and elementary bodies (2), as demonstrated by the use of Morosow's technique, permitted an identification of the constituents of the two boundaries; collodion particles coated with rabbit serum protein took a tan color in contrast to the jet black displayed by elementary bodies. Parts a and b of Fig.…”

Micro-Electrophoresis of Cowpox and Vaccinia Viruses in Molar Sucrose

“…Thus Shedlovsky & Smadel (1940) showed that under carefully controlled conditions, purified preparations of vaccinia virus migrated as single boundaries in the Tiselius cell. Similar experiments by McFarlane (1940) were hampered by endo-osmotic disturbances, a difficulty which Shedlovsky & Smadel were able to eliminate by the addition of a small amount of protein to their suspensions.…”