Monoclonal Antibodies to Guinea-pig Cytomegalovirus: An Immunoelectron Microscopic Study

Abstract

SUMMARYThree groups of monoclonal antibodies which reacted with cells infected by guineapig cytomegalovirus (GPCMV) were prepared. The first group of antibodies immunoprecipitated a 50 000 mol. wt. (50K) polypeptide of GPCMV-infected cells. This polypeptide was identified as part of the nuclear inclusion by immunofluorescence. This nuclear fluorescence was markedly diminished when the infected cells were incubated in the presence of phosphonoacetic acid. By immunoelectron microscopy the antibodies reacted mainly with filamentous structures (26 to 28 nm in diameter) in nuclear inclusions and occasionally stained nucleocapsids. Neither intracytoplasmic nor extracellular virions reacted with the antibodies. Therefore, the 50K protein with which the monoclonal antibodies reacted was nuclear inclusion-specific and a nonstructural protein. The second group of antibodies reacted with a 76K polypeptide of the infected cells which was a matrix protein found in both cytoplasmic inclusions and extracellular dense virions. The third group of antibodies mainly reacted with a virion core protein by immunoelectron microscopy.

Subcellular distribution of the major immediate early proteins of human cytomegalovirus changes during infection

“…Although the biological significance of the redistribution of IEt proteins during HCMV infection is not known, in the late phase of infection these proteins may play some role other than in gene regulation. Nucleocapsids and cores in the nuclear inclusions were sometimes stained by either MAb e-8 or MAb E3; similar findings have been obtained using cells infected with guinea-pig CMV using an MAb specific for the 50K DNA-binding protein (Tsutsui et al, 1986;Nogami-Satake & Tsutsui, 1988). It may be possible that a small amount of virus-encoded nuclear protein can be incorporated into the nucleocapsids by chance during viral morphogenesis.…”

“…), the cell monolayers were washed with phosphate-buffered saline (PBS) and fixed in periodatelysine-paraformaldehyde, as described by McLean & Nakane (1974), with 3% paraformaldehyde. Immunoelectron microscopy procedures were carried out basically as described previously (Tsutsui et al, 1986). Briefly, the cell monolayers were incubated in 10%, 15% and 20% sucrose in PBS, frozen in solid carbon dioxideethanol for 3s and thawed in 20% sucrose in PBS.…”

“…After polymerization the capsules and cells were detached from the 0000-9894 © 1991 SGM glass slides by heating on a hot plate at 80 °C for 5 min. Ultrathin section and electron microscopy were performed as described previously (Tsutsui et al, 1986).…”

Pathogenesis of cytomegalovirus-associated labyrinthitis in a guinea pig model

“…A mouse MAb to GPCMV was developed as described previously [24]. Briefly, six-week-old BALB/c mice were immunized with GPCMV strain 22122-infected GP embryonic cells.…”

Immunoblot analysis of the humoral immune response to cytomegalovirus in non-pregnant and pregnant guinea pigs

“…Guinea pig CMV proteins, like those of human CMV, appear to be synthetized in infected cells in a regulated cascade of immediate early, early, and late viral proteins [15]. To date, only one guinea pig CMV protein, a non structural DNA binding protein with a molecular weight of 50 kDa has been characterized [20,22]. Further, the immunogenic proteins of guinea pig CMV have not been identified.…”