Direct Functional Comparison of Murine Macrophages: BMDM, RAW 264.7 and J774A.1
Location
Room 232, Schewel Hall
Access Type
Campus Access Only
Presentation Type
Oral presentation
Entry Number
2437
Start Date
4-16-2025 9:15 AM
End Date
4-16-2025 9:30 AM
School
School of Medicine and Health Sciences
Department
Biomedical Science
Keywords
Macrophages, Inflammation, Nitric oxide, Bone marrow, Tumor necrosis factor, RAW 264.7, J774A.1
Abstract
This study conducts a comprehensive functional comparison between the widely used RAW 264.7 murine macrophage cell line, J774A.1 macrophage cell line and primary bone marrow-derived macrophages (BMDMs). Macrophages play essential roles in innate immunity, inflammation regulation, antigen presentation, and tissue homeostasis, making them critical subjects in immunological research. While RAW 264.7 cells offer experimental advantages in accessibility and reproducibility, previous research has identified significant differences between immortalized lines and primary cells in cytokine production profiles and inflammatory responses. This investigation addresses a fundamental methodological question in immunology research by directly comparing canonical macrophage functions—including nitric oxide production, TNF-α secretion, and iNOS expression—following lipopolysaccharide stimulation. Our experimental design included a RAW 264.7 LPS dose-response assay measuring nitric oxide production at various LPS concentrations (0, 1, 10, and 100 ng/mL). Macrophage function is evaluated by nitric oxide production using the Greiss reaction. For total nitrite RAW 264.7 and J774A.1 cells respond at equivalent levels, while BMDM appears to respond less vigorously. When response is adjusted to nitrite levels per 10,000 cells the RAW 264.7 cells have a greater response than either J774A.1 or BMDM. These differences could be due to two factors. Variability in generating accurate cell counts following experimental treatment with LPS is one problem. Additionally, metabolic activity in these different cell types could account for differences in response depending upon how that response is quantified. More accurate measures of cell counts are necessary to determine if the differences we observed are real or just a remnant of cell counting variability.
Primary Faculty Mentor(s)
Dr. David O. Freier
Primary Faculty Mentor(s) Department
Biology
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Direct Functional Comparison of Murine Macrophages: BMDM, RAW 264.7 and J774A.1
Room 232, Schewel Hall
This study conducts a comprehensive functional comparison between the widely used RAW 264.7 murine macrophage cell line, J774A.1 macrophage cell line and primary bone marrow-derived macrophages (BMDMs). Macrophages play essential roles in innate immunity, inflammation regulation, antigen presentation, and tissue homeostasis, making them critical subjects in immunological research. While RAW 264.7 cells offer experimental advantages in accessibility and reproducibility, previous research has identified significant differences between immortalized lines and primary cells in cytokine production profiles and inflammatory responses. This investigation addresses a fundamental methodological question in immunology research by directly comparing canonical macrophage functions—including nitric oxide production, TNF-α secretion, and iNOS expression—following lipopolysaccharide stimulation. Our experimental design included a RAW 264.7 LPS dose-response assay measuring nitric oxide production at various LPS concentrations (0, 1, 10, and 100 ng/mL). Macrophage function is evaluated by nitric oxide production using the Greiss reaction. For total nitrite RAW 264.7 and J774A.1 cells respond at equivalent levels, while BMDM appears to respond less vigorously. When response is adjusted to nitrite levels per 10,000 cells the RAW 264.7 cells have a greater response than either J774A.1 or BMDM. These differences could be due to two factors. Variability in generating accurate cell counts following experimental treatment with LPS is one problem. Additionally, metabolic activity in these different cell types could account for differences in response depending upon how that response is quantified. More accurate measures of cell counts are necessary to determine if the differences we observed are real or just a remnant of cell counting variability.