Feucht H E, Hadam M R, Frank F, Riethmüller G
J Immunol Methods. 1980;38(1-2):43-51. doi: 10.1016/0022-1759(80)90329-4.
In a comparative study, human peripheral T lymphocytes were separated as E rosettes by density centrifugation through various gradient media. Sheep red blood cells (SRBC) were removed by dissociation of the E rosettes at 37 degrees C with subsequent centrifugation on a similar density gradient prewarmed to 37 degrees C. In particular, gradients made of Ficoll Urovison were compared with Percoll gradients with regard to both separation steps. Using Percoll gradients, a maximal T cell recovery of 75% was obtained, whereas Ficoll separation yielded only 46%. T lymphocytes separated with Percoll exhibited equal viability compared to Ficoll isolated cells and consisted of 98% EAET-RFC. No inhibition of cellular function by Percoll treatment was detected, whereas Ficoll treatment led to an impaired mitogenic response. An inherent mitogenicity of Percoll was not observed. The method described results in considerably shortened centrifugation times due to the low viscosity of the Percoll medium and simultaneously seems to be less harmful to the rather fragile rosettes. Reproducibility was found to depend on careful control of density and osmolarity of the Percoll medium.
在一项对比研究中，通过在各种梯度介质中进行密度离心，将人外周血T淋巴细胞分离为E花环。通过在37℃解离E花环，随后在预热至37℃的类似密度梯度上进行离心，去除绵羊红细胞（SRBC）。特别地，就两个分离步骤而言，比较了由Ficoll Urovison制成的梯度与Percoll梯度。使用Percoll梯度时，T细胞最大回收率为75%，而Ficoll分离仅产生46%。与Ficoll分离的细胞相比，用Percoll分离的T淋巴细胞具有同等的活力，并且由98%的EAET-RFC组成。未检测到Percoll处理对细胞功能的抑制作用，而Ficoll处理导致有丝分裂反应受损。未观察到Percoll具有内在的促有丝分裂活性。由于Percoll介质的低粘度，所述方法导致离心时间显著缩短，并且同时似乎对相当脆弱的花环危害较小。发现重现性取决于对Percoll介质的密度和渗透压的仔细控制。
