Why is it important to pH the assay medium being used?

Seahorse XF assays are designed to measure changes in metabolic analytes (i.e. dissolved oxygen and pH), which is reflected in the oxygen consumption (OCR) and extracellular acidification rates (ECAR). Additionally, the cells being tested are acclimatized to a neutral pH environment- additional changes to their pH can affect their metabolic profile/ phenotype. Seahorse recommends adjusting the pH of the final assay media which will include all media supplements to 7.4 at 37°C prior to running the assay.