rM-CSF efficiently replaces L929 in generating mouse and rat bone marrow-derived macrophages for in vitro functional studies of immunity to intracellular bacteria.

rM-CSF efficiently replaces L929 in generating mouse and rat bone marrow-derived macrophages for in vitro functional studies of immunity to intracellular bacteria. J Immunol Methods. 2019 Nov 02;:112693 Authors: Rice HM, Rossi AP, Bradford MK, Elkins KL, De Pascalis R Abstract Methods used to prepare bone marrow-derived macrophages (BMDMs) may influence the outcomes of immunological assays in which they are used. Supernatant conditioned by growth of L929 cells has often been used to generate mouse macrophages from bone marrow in vitro but is subject to lot-to-lot variability. To reduce experimental variability and to standardize techniques across laboratories, we investigated recombinant M-CSF (rM-CSF) as an alternative supplement for BMDM maturation in the context of macrophage infection, using the intracellular bacterium Live Vaccine Strain (LVS) of Francisella tularensis as a prototype. We compared rM-CSF with L929 supernatant in terms of their effects on mouse and rat macrophage growth, maturation patterns, surface marker expression, and the expression of selected genes. Further, we compared macrophage infectivity and bacterial replication using LVS. Finally, we compared the in vitro function of BMDMs co-cultured with splenocytes from vaccinated animals in terms of their control of intramacrophage bacterial replication, as well as production of cytokines and nitric oxide. We demonstrated that rM-CSF produced BMDMs with similar, o...
Source: Journal of Immunological Methods - Category: Allergy & Immunology Authors: Tags: J Immunol Methods Source Type: research