Dendritic Cell Generation from Mouse Bone Marrow

Were isolated from mouse bone marrow and resuspended in a complete medium containing RPMI 1640, 10% FBS, and 1% penicillin/streptomycin. The cell concentration was adjusted to 0.5-1 × 10⁶/ml, and the cells were seeded into a 24-well culture plate. Recombinant mouse GM-CSF (20ng/ml, PeproTech Inc., #315-03, USA) and IL-4 (10ng/ml, MCE, #HY-P701093) were added on day 0 of culture. On day 2, the culture plate was gently shaken and 3/4 of the culture medium was replaced with fresh medium supplemented with cytokines. These steps were repeated every two days. On the 6th day, 1ug/ml of LPS (Sigma, #L2880) was added to the medium. On the 8th day, suspended and loosely adherent cells were collected as mature dendritic cells.

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Xu Q., Hua X., Li B., Jiang B., Jin J., Wu R., Gu Y., Xu H., Cheng Q., Zhu S., Zhang F., Lv T, & Song Y. (2025). Intrinsic STING of CD8 + T cells regulates self-metabolic reprogramming and memory to exert anti-tumor effects. Cell Communication and Signaling : CCS, 23, 99.

Publication 2025

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Variable analysis

independent variables

Recombinant mouse GM-CSF (20ng/ml)
Recombinant mouse IL-4 (10ng/ml)
LPS (1ug/ml)

dependent variables

Mature dendritic cells

control variables

RPMI 1640 medium
10% FBS
1% penicillin/streptomycin
Cell concentration (0.5-1 × 10^6/ml)
Culture plate (24-well)

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

Annotations

Based on most similar protocols

To optimize experimental performance, the protocols recommend using GM-CSF (20-40 ng/ml) and IL-4 (10-1000 U/ml) for DC differentiation (Protocols 1-5).

The protocols use LPS (1 μg/ml) as a maturation stimulus for DCs on day 6 or day 5 to generate mature DCs (Input Protocol, Protocol 4, Protocol 5).

The protocols use 10% FBS in the culture medium to support DC differentiation and maturation (Input Protocol, Protocols 1-5).

The protocols recommend adjusting the cell concentration to 2 × 10^5 cells/ml or 0.5-1 × 10^6 cells/ml for optimal DC generation (Protocols 1-3, Input Protocol).

The protocols suggest flushing the bone marrow from the long bones and tibias, and removing cell clumps by passing the cells through a 70 μm cell strainer (Input Protocol, Protocols 1-3).

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