Roswell Park Memorial Institute (RPMI) 1640 medium was originally developed to culture human leukemic cells in suspension and as a monolayer. RPMI 1640 has since been found suitable for a variety of mammalian cells including HeLa, Jurkat, MCF-7, PC12, PBMC, astrocytes and carcinomas.
RPMI 1640 is unique from other media because it contains the reducing agent glutathione and high concentrations of vitamins. RPMI 1640 contains biotin, vitamin B₁₂ and PABA which are not found in Eagle's Minimal Essential Medium or Dulbecco's Modified Eagle Medium. In addition, the vitamins inositol and choline are present in very high concentrations.

RPMI 1640 contains no proteins, lipids, or growth factors. Therefore, RPMI 1640 requires supplementation, commonly with 10% Fetal Bovine Serum (FBS). RPMI 1640 uses a sodium bicarbonate buffer system (2.0 g ⁄ L) and therefore requires a 5-10% CO₂ environment to maintain physiological pH.

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