Rapid single-step method for purification of polymorphonuclear leucocytes from blood of patients with rheumatoid...

Previous studies have shown that blood samples from healthy subjects and patients with a variety of different diseases separate into distinct mononuclear leucocyte (MNL) and polymorphonuclear leucocyte (PMNL) bands when centrifuged on a Ficoll-Hypaque medium of density 1.114 g/ml. Results from the present study showed that blood samples from patients with rheumatoid arthritis (RA) failed to separate into discrete cellular fractions under the same conditions. In particular, the PMNL were not recoverable because the erythrocytes only partially sedimented. However, following modification of the separation medium involving (a) reduction in the final Ficoll concentration from 8% to 7%, and (b) an alteration in the ratio of meglumine Hypaque:sodium Hypaque from 5.7:2.8 to 2.0:7.5, a clear band of PMNL of high purity could be obtained with blood samples from patients with RA. The modification of this previously described, simple, brief, one-step, density gradient centrifugation method achieved separation of peripheral blood PMNL required for studies of leucocyte function in patients with RA.