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Proteomics characterisation of the L929 cell supernatant and its role in BMDM differentiation

Lookup NU author(s): Rachel Heap, Dr Jose Luis Marin-RubioORCiD, Dr Julien Peltier, Dr Tiaan Heunis, Dr Abeer Dannoura, Adam Moore, Professor Matthias TrostORCiD



This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


© 2021 Heap et al.BMDMs are a key model system to study macrophage biology in vitro. Commonly used methods to differentiate macrophages from BM are treatment with either recombinant M-CSF or the supernatant of L929 cells, which secrete M-CSF. However, little is known about the composition of L929 cell-conditioned media (LCCM) and how it affects the BMDM phenotype. Here, we used quantitative mass spectrometry to characterise the kinetics of protein secretion from L929 cells over a 2-wk period, identifying 2,193 proteins. Whereas M-CSF is very abundant in LCCM, we identified several other immune-regulatory proteins such as macrophage migration inhibitory factor (MIF), osteopontin, and chemokines such as Ccl2 and Ccl7 at surprisingly high abundance levels. We therefore further characterised the proteomes of BMDMs after differentiation with M-CSF, M-CSF + MIF, or LCCM, respectively. Interestingly, macrophages differentiated with LCCM induced a stronger anti-inflammatory M1 phenotype that those differentiated with M-CSF. This resource will be valuable to all researchers using LCCM for the differentiation of BMDMs.

Publication metadata

Author(s): Heap RE, Marin-Rubio JL, Peltier J, Heunis T, Dannoura A, Moore A, Trost M

Publication type: Article

Publication status: Published

Journal: Life Science Alliance

Year: 2021

Volume: 4

Issue: 6

Print publication date: 01/06/2021

Online publication date: 14/04/2021

Acceptance date: 07/04/2021

Date deposited: 05/05/2021

ISSN (electronic): 2575-1077

Publisher: Life Science Alliance LLC


DOI: 10.26508/lsa.202000957

PubMed id: 33853969


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Funder referenceFunder name
215542/Z/19/ZWellcome Trust