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Published on: March 25, 2016
An optimized method to differentiate HL60 cells into neutrophil-like cells
Samuel P Collie1, Grant Yu2, Suraj S Rawat3
1Cellular and Molecular Biology Program, University of Michigan Medical School, 1135 Catherine St., Ann Arbor, MI 48109, USA.
Journal of Leukocyte Biology
|June 22, 2026
Summary
A new protocol using sequential all-trans retinoic acid (ATRA) and dimethyl sulfoxide (DMSO) treatments differentiates HL60 cells into neutrophil-like cells. These cells exhibit improved morphology and functionality, closely mimicking mature human neutrophils.
Area of Science:
- Cell Biology
- Hematology
- Immunology
Background:
- The HL60 promyelocytic leukemia cell line is a valuable model for studying neutrophil biology.
- Standard differentiation methods using all-trans retinoic acid (ATRA) or dimethyl sulfoxide (DMSO) yield cells with incomplete mature granulocyte features.
Purpose of the Study:
- To develop an improved differentiation protocol for HL60 cells that generates neutrophil-like cells with enhanced morphology and function.
- To overcome the limitations of existing ATRA and DMSO differentiation methods.
Main Methods:
- A sequential differentiation protocol involving one day of ATRA treatment followed by four days of DMSO treatment (A1D4) was developed.
- Morphological analysis, including nuclear segmentation and lamin A/C expression, was performed.
- Functional assays assessing chemotaxis, oxidative burst, and phagocytosis were conducted.
Main Results:
- A1D4-differentiated HL60 cells displayed segmented nuclei and reduced lamin A/C expression, characteristic of mature neutrophils.
- These cells exhibited robust chemotactic, oxidative burst, and phagocytic activities.
- The A1D4 protocol successfully combined the benefits of ATRA and DMSO treatments.
Conclusions:
- The sequential A1D4 differentiation protocol effectively produces HL60-derived neutrophil-like cells that more accurately mimic the phenotype and function of primary human neutrophils.
- This improved model system offers enhanced capabilities for research in neutrophil biology and related fields.
