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 T-cells enrichment from the spleen of a mouse by negative selection using a magnetic separator. The technique involves targeting the other cells in the sample for depletion using antibodies or ligands directed against specific cell surface antigens.
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Day 1: Prepare T-cells from the donor mice
The authors have nothing to disclose.
RPMI 1640 | Thermo Fisher Scienctific | 11875-093 | Media |
MidiMACS | Miltenyi Biotec | 130-042-302 | T-cell enrichment |
0.5 M EDTA pH 8.0 100ML | Fisher Scientific | BP2482100 | MACS buffer |
10X PBS | Fisher Scientific | BP3994 | MACS buffer |
Anti-Biotin MicroBeads | Miltenyi Biotec | 130-090-485 | T-cell enrichment |
Anti-Human/Mouse CD45R (B220) | Thermo Fisher Scientific | 13-0452-85 | T-cell enrichment |
Anti-Mouse CD4 Biotin | Thermo Fisher Scientific | 13-0041-86 | T-cell enrichment |
CD11b | Thermo Fisher Scientific | 13-0112-85 | T-cell enrichment |
CD25-biotin | Thermo Fisher Scientific | 13-0251-82 | T-cell enrichment |
CD45R | Thermo Fisher Scientific | 13-0452-82 | T-cell enrichment |
CD49b Monoclonal Antibody (DX5)-biotin | Thermo Fisher Scientific | 13-5971-82 | T-cell enrichment |
Anti-Mouse TER-119 Biotin | Thermo Fisher Scientific | 13-5921-85 | T-cell enrichment |
Anti-mouse CD45.1 PE | Thermo Fisher Scientific | 12-0900-83 | Flow cytometry analysis |
C57BL/6 mice | Charles River | 27 | Donors/Recipients |
Flow cytometry tubes | Fisher Scientific | 352008 | Flow cytometry analysis |
Cell strainer 40 uM | Thermo Fisher Scientific | 22363547 | Cell preparation |