A rapid and simple method for the separation of pure lymphocytes from horse blood

A method for the separation of pure and viable lymphocytes and granulocytes from the same blood sample in horses was reported. By centrifuging equine heparinized blood at 100 xg for 10 min at room temperature (r.t.), the resulting supernatant plasma was an almost pure (97.71 +/- 0.30%; n = 15) suspension of highly viable (98.72 +/- 0.28%) lymphocytes. When sodium citrate was used as an anticoagulant, lymphocyte suspensions collected in the same manner showed lower purity (87.89 +/- 1.59%; n = 9) and higher yields (56.56 +/- 3.89%, n = 9 versus 36.11 +/- 2.23%, n = 15). Where needed, a further centrifugation at 250 xg for 3 min (r.t.) of heparinized lymphocyte preparations removed an average of 87.39% (n = 15) contaminating platelets. A suspension of 85.96 +/- 2.20% pure granulocytes (93.23 +/- 1.74% neutrophils; n = 14) with minimal contamination by erythrocytes and high viability (93.11 +/- 1.26%) was obtained by performing a flash red blood cell lysis on the white-greyish layer resulting from the centrifugation of the heparinized blood samples. Among the several methods available, the procedure described herein is easy, rapid, cheap and reproducible.