Isolating Cord Blood Hematopoietic Stem Cells to Generate Human Immune System Mice

Begin with human umbilical cord blood cells, including CD34⁺ hematopoietic stem cells, or HSCs.

Add antibodies to block cellular Fc receptors, preventing non-specific binding of HSC-specific magnetic beads.

Introduce the antibody-coated magnetic beads targeting CD34⁺ HSCs. Centrifuge and discard the supernatant.

Resuspend the cells in a buffer. Pass them through a magnetic separator to retain bead-bound HSCs; unbound cells flow through.

Remove the magnetic field to collect the HSCs.

Centrifuge and resuspend the cells in a growth medium. Plate the cells and incubate.

The medium contains growth factors and cytokines that promote HSC survival and expansion.

Harvest the cells. Add a freezing medium, transfer to a cryovial, and freeze.

To generate a human immune system, or HIS, mouse model, thaw the HSCs. Add them to a pre-warmed medium.

Centrifuge and resuspend the cells in a buffer.

The cells are ready for injection into an immunodeficient mouse pup for HIS model generation.

To begin, resuspend the CD34-positive cell pellet isolated from cord blood in 300 microliters of magnetic cell separator buffer per 1 x 10⁸ cells. Add 100 microliters of FcR blocking reagent per 1 x 10⁸ cells, fol