Negative Immunomagnetic Selection: A Method to Purify B-cells from Peripheral Blood Mononuclear Cells

Peripheral blood mononuclear cells or PBMCs, include B cells, T cells, natural killer or NK cells, and monocytes. B cells produce immunoglobulin molecules to fight infection. In leukemia, these cells multiply and metastasize to different organs.

To isolate B cells, begin with a suspension of isolated PBMCs. Add primary antibody mixture directed against specific cell surface molecules called cluster of differentiation or CD. Specific antibodies bind to the CD-3 molecule on T cells, CD-14 molecule on monocytes, and CD-16 molecule on NK cells, leaving the B cells unlabeled.

Next, wash the sample to remove the unbound antibodies. Add secondary antibody-conjugated magnetic microbeads to the sample. Secondary antibodies bind to the FC region of primary antibodies attached to specific cells. Place the tube containing the labeled cell suspension under the influence of a magnetic field. In the field, the magnetically labeled cells migrate and bind to the tube wall, leading to the negative selection of the unwanted cells. Collect the supernatant containing enriched B cells into a fresh collection tube.

In this protocol, we will perform a negative immunomagnetic selection of leukemic B cells