Nanoparticle-Based Enrichment and Expansion of Antigen-Specific CD8+ T Cells
Published: August 31, 2023
Abstract
Source: Hickey, J. W., et al. Enrich and Expand Rare Antigen-specific T Cells with Magnetic Nanoparticles. J. Vis. Exp. (2018).
This video describes a protocol for isolating rare-antigen-specific CD8+ T cells using magnetic nanoparticle-based artificial antigen-presenting cells (aAPCs) functionalized with antigenic peptide-loaded MHC-Igs and anti-CD28 antibodies. Interaction of the aAPCs with CD8+ T cells specifically expressing the antigenic peptide-targeting T cell receptors, followed by their expansion in vitro, results in the enrichment and expansion of this CD8+ T cell population.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Enrich Antigen-specific CD8+ T Cells with Prepared Nanoparticle Artificial Antigen-Presenting Cells. Isolate CD8+ T cells. Euthanize the animals by exposure to isoflurane followed by cervical dislocation. Remove the spleen and lymph nodes from wild-type C57BL/6j mice and place the…
開示
The authors have nothing to disclose.
Materials
| DimerX I: Recombinant Soluble Dimeric Human HLA-A2:Ig Fusion Protein | BD Biosciences | 551263 | |
| DimerX I: Recombinant Soluble Dimeric Mouse H-2D[b]:Ig | BD Biosciences | 551323 | |
| DimerX I: Recombinant Soluble Dimeric Mouse H-2K[b]:Ig Fusion Protein | BD Biosciences | 550750 | |
| nanomag-D-spio, NH2, 100 nm nanoparticles | Micromod | 79-01-102 | |
| 96-Well Half-Area Microplate, black polystyrene | Corning | 3875 | |
| Super Mag NHS-Activated Beads, 0.2 µm | Ocean Nanotech | SN0200 | |
| Anti-Biotin MicroBeads UltraPure | Milteny | 130-105-637 | |
| EZ-Link NHS-Biotin | ThermoFisher | 20217 | |
| C57BL/6J (B6 wildtype) mice | Jackson Laboratory | 664 | |
| CD8a+ T Cell Isolation Kit, Mouse | Miltenyi | 130-104-075 | |
| MS Columns | Milteny | 130-042-201 | |
| LS Columns | Milteny | 130-042-401 | |
| Streptavidin-Phycoerythrin, SAvPE | Biolegend | 405203 | |
| N52 disk magnets of 0.75 inches | K&J Magnetics | DX8C-N52 | |
| APC anti-mouse CD8a Antibody, clone 53-6.7 | Biolegend | 100711 | |
| LIVE/DEAD Fixable Green Dead Cell Stain Kit, for 488 nm excitation | ThermoFisher | L-34969 |
