Use of Methyl Cellulose Gel as a Substitute for Agar in Tissue-Culture Overlays

Abstract

IN the course of experiments designed to assay non-cytopathogenic animal viruses by Dulbecco's plaque-count method¹ using fluorescent antibody² to identify the plaques, a non-fluorescent substitute was required for the agar used in the overlay. Methyl cellulose has been used by Earle³ to increase the viscosity of tissue-culture fluids for suspended-cell cultures, and appears to be harmless to animal tissue. An investigation of its properties showed it to be capable of forming highly viscous aqueous solutions which would gel spontaneously on raising their temperature. The presence of salts at physiological concentrations lowers the temperature of gelation. Thus one can make a solution of methyl cellulose containing tissue nutrients which, after being added as a liquid at room temperature to the tissue, will set quite firmly soon after being placed in an incubator at 37° C. A further advantage is that this overlay can be removed entirely without disturbing the tissue; this is achieved by cooling the culture in a refrigerator for a short time, during which the gel liquefies, enabling it to be removed by pipetting.