This protocol is intended for positive magnetic isolation or depletion of CD25high T cells from human samples such as mononuclear cells (MNCs) or whole blood. Dynabeads® bind to the target cells during a short incubation, and then the bead-bound cells are separated by a magnet.
Depletion – discard the bead-bound cells and use the remaining, untouched cells for any application.
Positive isolation – discard the supernatant and use the bead-bound cells for downstream molecular applications.
This protocol describes isolation from up to 2.5x107 MNCs. Isolated cells are ideal for use in downstream molecular assays such as protein and gene expression analysis.
Isolated cells may be lysed while attached to the beads and directly processed for further molecular analysis. For recommended products and protocols visit the Immunology Research Guide page.
Approximately 1x106 T cells are present per ml human blood, and about 5–10% of these T cells strongly express the CD25 antigen. This protocol describes depletion or positive isolation of CD25high T cells from 2.5x107 human mononuclear cells (MNCs) using Dynabeads® CD25.
For scale-up to larger cell numbers, adjust the volumes accordingly.
Dynabeads® Washing Procedure
Dynabeads® should be washed before use.
Depletion or Positive Isolation of CD25+ cells
For rapid and consistent results in protein or gene expression analysis, lyse the CD25+ cells while they are still attached to the beads and directly process for further molecular analysis.For further technical advice please visit Pan Mouse IgG - Depletion or Positive Isolation of Cells from Different Species.
Invitrogen Dynal® AS complies with the Quality System Standards ISO 9001:2000 and ISO 13485:2003.
This product is stable until the expiry date stated on the label when stored unopened at 2-8°C. Store opened vials at 2-8°C and avoid bacterial contamination. Keep Dynabeads® in liquid suspension during storage and all handling steps, as drying will result in reduced performance. Resuspend well before use.
Warnings And Limitations
This product is for research use only. Not intended for any animal or human therapeutic or diagnostic use unless otherwise stated. Follow appropriate laboratory guidelines. This product contains 0.02% sodium azide as a preservative, which is cytotoxic.
Avoid pipetting by mouth!
Sodium azide may react with lead and copper plumbing to form highly explosive metal azides. When disposing through plumbing drains, flush with large volumes of water to prevent azide build up. Certificate of Analysis (CoA) is available upon request. Material Safety Data Sheet (MSDS) is available at .
PBS (phosphate buffered saline) supplemented with 0.1% BSA and 2mM EDTA, e.g. Gibco cat.no. 14190-094.
If preferred, PBS with 2% fetal calf serum and 1 mM EDTA may be used.
Preparation of MNC from Buffy Coat
This method gives low platelet numbers and is recommended for use with Dynabeads® products.
For starting samples other than buffy coat please see the page Pan Mouse IgG - Depletion or Positive Isolation of Cells from Different Species for recommended MNC preparation procedures. Please contact Invitrogen Dynal® for further technical information (see contact details).
For Research Use Only. Not for use in diagnostic procedures.