A Comparison of Function in RAW 264.7 Murine Macrophages Grown in DMEM and RPMI 1640
Location
Room 232, Schewel Hall
Access Type
Campus Access Only
Start Date
4-17-2024 8:30 AM
End Date
4-17-2024 8:45 AM
College
Lynchburg College of Arts and Sciences
Department
Biology
Abstract
The RAW 264.7 murine macrophage is commonly used for the study of inflammation and a common cell line in mammalian cell culture. The standard medium for its growth and maintenance is Dulbecco's Modified Eagle's Medium (DMEM), but in a portion of published work, RPMI 1640, is substituted. RPMI 1640 lacks the high level of glucose present in DMEM (4.5 g/L). The purpose of this work compares RAW 264.7 cells grown and maintained in DMEM and RPMI 1640. Both media are supplemented with 10% fetal bovine serum, sodium bicarbonate, and 100 IU Penicillin Streptomycin 100μg. DMEM includes the addition of 4.5 g/L glucose. Following a dose response exposure of RAW 264.7 to a range of 0 ng/mL to 100 ng/mL lipopolysaccharide (LPS) from E. coli (serotype O55:B5) for 24 hours in a 5%CO2/ 37°C environment, nitrite is measured. Cells are maintained in continuous culture with passage every 3 days. Experimentation is carried out on a 24-well plate 500μl volume at 4x105 cells per well. After 24-hour exposure to LPS, the Greiss reaction is used to measure nitric oxide response in supernatants. Initial results suggest RAW 264.7 cells grown in RPMI 1640 have a decreased nitric oxide response. Once the difference between DMEM and RPMI 1640 is established, addition of glucose to RPMI 1640 will be done to rescue RAW 264.7 in low-glucose RPMI 1640 to recover their nitric oxide response levels observed in DMEM cultures.
Faculty Mentor(s)
Dr. David Freier
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A Comparison of Function in RAW 264.7 Murine Macrophages Grown in DMEM and RPMI 1640
Room 232, Schewel Hall
The RAW 264.7 murine macrophage is commonly used for the study of inflammation and a common cell line in mammalian cell culture. The standard medium for its growth and maintenance is Dulbecco's Modified Eagle's Medium (DMEM), but in a portion of published work, RPMI 1640, is substituted. RPMI 1640 lacks the high level of glucose present in DMEM (4.5 g/L). The purpose of this work compares RAW 264.7 cells grown and maintained in DMEM and RPMI 1640. Both media are supplemented with 10% fetal bovine serum, sodium bicarbonate, and 100 IU Penicillin Streptomycin 100μg. DMEM includes the addition of 4.5 g/L glucose. Following a dose response exposure of RAW 264.7 to a range of 0 ng/mL to 100 ng/mL lipopolysaccharide (LPS) from E. coli (serotype O55:B5) for 24 hours in a 5%CO2/ 37°C environment, nitrite is measured. Cells are maintained in continuous culture with passage every 3 days. Experimentation is carried out on a 24-well plate 500μl volume at 4x105 cells per well. After 24-hour exposure to LPS, the Greiss reaction is used to measure nitric oxide response in supernatants. Initial results suggest RAW 264.7 cells grown in RPMI 1640 have a decreased nitric oxide response. Once the difference between DMEM and RPMI 1640 is established, addition of glucose to RPMI 1640 will be done to rescue RAW 264.7 in low-glucose RPMI 1640 to recover their nitric oxide response levels observed in DMEM cultures.