Bone Marrow-Derived Dendritic Cells Generation: A Method to Generate Dendritic Cells from Mouse Bone Marrow

Abstract

Source:Nguyen, H. D., et al. Bone Marrow Transplantation Platform to Investigate the Role of Dendritic Cells in Graft-versus-Host Disease. J. Vis. Exp.. (2020).

This video describes a protocol for the generation of dendritic cells--key antigen-presenting cells that activate T cells--from the bone marrow of an adult mouse.

Protocol

1. Generate bone marrow-derived dendritic cells (BM-DCs).

1. Isolate bone marrow from the femurs and tibias of WT or factor B (fB)^-/- mice on the B6 background. Briefly, collect the tibia and femur from both legs using forceps and scissors. Put them in ice-cold RPMI containing 1% FCS and 100 U/mL penicillin/streptomycin and 2 mM L-glutamine (1% RPMI) in 50 mL conical tubes. Clean the femur and the tibia bones thoroughly by removing all the muscle tissues using forceps and scissors. Transfer the femur and the tibia from the 50 mL conical tubes to a 92 mm diameter Petri dish containing 1% RPMI. Using scissors, cut the ends of the femur or tibia.....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
10X PBS	Fisher Scientific	BP3994	MACS buffer
B6 fB-/- mice	University of Central Florida	In house	Recipients
B6.Ly5.1 (CD45.1 + ) mice	Charles River	546	Donors
BALB/c mice	Charles River	28	Transplant recipients
C57BL/6 mice	Charles River	27	Donors/Recipients
Fetal Bovine Serum (FBS)	Atlanta Bilogicals R&D system	D17051	Cell Culture
Lipopolysaccharide (LPS)	Millipore Sigma	L4391-1MG	DC mature
New Brunswick Galaxy 170R incubator	Eppendorf	Galaxy 170 R	Cell Culture
Penicillin+streptomycinPenicillin/ Streptomycin (10,000 units penicillin / 10,000 mg/ml strep)	GIBCO	15140	Media
RPMI 1640	Thermo Fisher Scienctific	11875-093	Media