 |
 |
 |
 |
>I read about freezing cells for controls in texts of all sorts, but none
>describe a procedure for doing so. We want to freeze not only peripheral
>blood leukocytes, but also BM and tissues (mostly lymph nodes).
> If someone would be so kind as to send a reference to a procedure, or
attach
>
>a description of a procedure, I would be most appreciative.
We have had good luck freezing PBLs in 10% DMSO and complete medium (RPMI
1640 with 10-15% serum). Freezing rate is important however. Surround the
freezing vials with ~1cm of styrofoam (we use styrofoam trays from Corning or
Falcon 12x75 mm conical centrifuge tubes) and place them in a -80 C freezer
overnight, then transfer them to liquid nitrogen or a -135 C freezer. Thaw
rapidly in a 37 C water bath. With this procedure we typically achieve
2.5-3% C.V. for PI staining of hypotonically lysed cells.
Thomas L. Morgan, Ph.D.
Regional Research Lab
Kaiser Permanente Medical Center
1515 N. Vernont Ave
Los Angeles, CA 90027
|
|
|
|