A subscription to JoVE is required to view this content. Sign in or start your free trial.
Chapters
- 00:00Title
- 00:09Introduction
- 01:05T Cell Staining
- 03:21Reagents and Equipment
- 05:44Assemble Materials and Equipment at Workspace
- 06:04Step 1: PBMC Isolation
- 08:51Step 2: CD4+ Cell Separatioin
- 11:01Step 3: In Vitro Expansion Culture
- 13:34Step 4: Visualizing T Cells by Tetramer Staining
- 15:33Conclusion
This procedure demonstrates the purification and in vitro expansion of antigen specific CD4+ T cells from whole peripheral blood and their visualization using MHC class II tetramers. Tetramers permit the direct visualization of T cells with a single antigen specificity and defined MHC class II restriction.
Tags
CD4+ T CellsMHC Class II TetramersAntigen SpecificFlow CytometryPeptide Loaded MHCT-cell ReceptorStreptavidinStaining ReagentsIn Vitro AmplificationMononuclear CellsPeripheral BloodFicoll UnderlayNegative SelectionBiotinylated Antibody CocktailMagnetic BeadsAdherent CellsAntigen Presenting CellsIL-2Surface AntibodiesFlow Cytometry Analysis
