Rhinovirus and zinc part 3

So far in my experiments to understand inhibition of rhinovirus replication by ZnCl2 I’ve found that at a concentration of 0.1 mM, viral plaque formation is inhibited but not sufficiently to be able to select resistant mutants. Attempts to use higher concentrations of the metal have consistently failed.

When I initially I tried higher concentrations of ZnCl2 in the plaque assay (0.2, 0.3, 0.4 mM) the cell monolayers looked poor. I thought one reason for this apparent toxicity was that the HeLa cell monolayers were too sparse. Last week I repeated the experiment using plates of HeLa cells seeded with 2 or 2.25 million cells each the night before. I infected the cells with two different amounts of rhinovirus type 1a, added a semisolid overlay with or without ZnCl2, and incubated for four days. The results are shown below.

rhinovirus_zinc_3

It’s clear that seeding the plates with 2 or 2.25 million HeLa cells made no difference: 0.2 mM ZnCl2 is still toxic to the monolayers. I find it surprising that the cells have no problem with 0.1 mM ZnCl2 but fare poorly at twice the concentration; but that is the observation and I’ll have to work around it.

Tomorrow’s experiment: try intermediate concentrations of ZnCl2 in the plaque assay, between 0.1 and 0.2 mM. It might be possible to identify a concentration that does not harm the cells but inhibits viral plaque formation >99%.