25mM HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid ) pH7 solution buffers are a media with a zwitterion organic chemical buffering agent; one of the twenty Good's buffers. HEPES DMEM medium is widely used in cell culture, largely because it is better at maintaining physiological pH 7 tot 7,5 despite changes in carbon dioxide concentration (produced by cellular respiration) when compared to bicarbonate buffers, which are also commonly used in cell culture. The dissociation of water decreases with falling temperature, but the dissociation constants (pK) of many other buffers do not change much with temperature. HEPES, 25 mM in RPMI and DMEM is like water in that its dissociation decreases as the temperature decreases. This makes HEPES RPMI 25 milli-molar media with or without L-glutamine a more effective buffering agent for maintaining enzyme structure and function at low temperatures.