CSF1 (M-CSF) is ubiquitously expressed and is the primary ligand for monocyte/macrophage homeostasis. IL-34 has restricted expression — mainly in the skin (keratinocytes) and brain (neurons) — and preferentially maintains Langerhans cells and microglia. Both bind CSF1R but at non-overlapping sites, and they share no sequence homology. Anti-CSF1R antibodies block both ligands, while anti-CSF1 antibodies block only CSF1, leaving IL-34-dependent populations (microglia, Langerhans cells) intact. This distinction has safety implications for therapeutic strategies.

Both, depending on the approach and context. Complete CSF1R blockade (high-dose small-molecule inhibitor or depleting anti-CSF1R antibody) eliminates macrophages by removing their survival signal. Partial or transient blockade can reprogram TAMs from an immunosuppressive M2 phenotype toward a pro-inflammatory M1 phenotype without full depletion. In preclinical tumor models, combination with anti-PD-1 works better with reprogramming than depletion, as residual pro-inflammatory macrophages can support anti-tumor immunity.

Isolate CD14⁺ monocytes from PBMCs by magnetic bead selection. Culture in RPMI + 10% FBS supplemented with 25-50 ng/mL recombinant M-CSF (CSF1) for 7 days, refreshing medium and cytokine on day 3-4. This produces M0 (unpolarized) macrophages. For M1 polarization, add IFN-γ + LPS on day 6. For M2 polarization, add IL-4 + IL-13. Verify differentiation by flow cytometry: CD14⁺CD68⁺CD163⁻ (M1) or CD14⁺CD68⁺CD163⁺CD206⁺ (M2).

Microglia are the brain's resident macrophages and depend on CSF1R signaling (primarily via IL-34 in the CNS) for survival. CSF1R inhibitor-mediated microglial depletion (using PLX5622 or BLZ945 in preclinical models) is a widely used tool for studying microglial contributions to Alzheimer's disease, ALS, traumatic brain injury, and glioblastoma. Upon inhibitor withdrawal, microglia rapidly repopulate from surviving progenitors. In glioblastoma, CSF1R blockade reprograms tumor-associated microglia/macrophages and synergizes with anti-PD-1 therapy.

Culture mouse bone marrow cells or human CD14⁺ monocytes with M-CSF (25 ng/mL) + RANKL (50-100 ng/mL) for 5-7 days. M-CSF provides survival signaling through CSF1R, while RANKL drives osteoclast commitment and fusion. Mature osteoclasts are identified as large multinucleated (3+ nuclei) TRAP-positive cells. Functional resorption can be assessed on dentine discs or calcium phosphate-coated plates. Anti-CSF1R blocking antibodies should inhibit both M-CSF-dependent survival and osteoclast formation.