Enhancement by calcium of the detection of cytomegalovirus in cells treated with dexamethasone and dimethyl sulfoxide

Abstract

MRC-5 cultures treated with dexamethasone, dimethyl sulfoxide, and calcium were compared with untreated cultures and cultures treated with dexamethasone and dimethyl sulfoxide for the detection of human cytomegalovirus by the shell vial method. The addition of calcium resulted in significantly increased growth of human cytomegalovirus AD169. Protein studies and cell counts showed renewed cell proliferation in the treated cultures. When low-titer clinical specimens were retested, 27.5% of all positives revealed were found only on the vials treated with calcium. In a group of freshly tested specimens, the calcium-treated group accounted for 20% of the positives.

Enhanced detection of cytomegalovirus in MRC-5 shell vials

“…Sensitivity can be increased even further by the inclusion of 2 mM calcium in the medium in the latter two steps (9). MRC-5 cells grown in shell vials from Whittaker Bioproducts were used in those experiments (8,9). Similar results were seen by investigators using MRC-5 cells grown in their own laboratory and treated with DEX alone (7).…”

“…Studies in our laboratory have shown that this sensitivity can be increased by pretreatment of the cells with medium containing dexamethasone (DEX) and by the addition of DEX plus dimethyl sulfoxide (DMSO) to the centrifugation and incubation steps (8). Sensitivity can be increased even further by the inclusion of 2 mM calcium in the medium in the latter two steps (9). MRC-5 cells grown in shell vials from Whittaker Bioproducts were used in those experiments (8,9).…”

“…In a previous study, we found that the cell treatments increased the protein content of individual vials, the result of * Corresponding author. a reproductive spurt in the static cultures (9). Since HCMV replicates best in actively metabolizing cultures, it would be reasonable to assume that the initiation of this reproductive spurt is at least partly responsible for the enhanced viral detection.…”

Induction of latent human cytomegalovirus by conventional gamma irradiation and prevention by treatment with INACTINE PEN110

“…MRC-5 cells were infected with CMV at a multiplicity of infection (MOI) of ≈ 0·02 for 2 h followed by the addition of MRC-5 growth medium supplemented with 2 m M calcium chloride (Sigma), 1% dimethylsulphoxide (DMSO) (Sigma) and 0·01 m M dexamethasone (Sigma) to enhance CMV replication [14]. The cells were collected 48 h postinfection by trypsinization, washed to remove extracellular virus and suspended in MEM.…”

“…MRC-5 cells, which are highly susceptible to spreading CMV infection [14], were used to model leucocytes actively replicating the virus. Real-time quantification of intracellular CMV DNA showed that gamma irradiation had no discernable effect on reducing the virus load immediately after treatment.…”