摘要
为探讨NK细胞克隆化培养的条件,我们首先将人外周血单个核细胞经rhIL-2诱导,使NK细胞得以扩增后,去除T细胞,得到相对纯化的NK细胞。经有限稀释,在饲养细胞及rhIL-2、PHA及LCM等培养条件下,获得NK细胞的单个克隆并进行鉴定。结果表明,每96孔板可获4-16个CD3^-CD56^+NK克隆,每个克隆的细胞数最多可达2.35×10~4,存活约3-5周。以丝裂霉素C(25μg/ml)作为饲养细胞抑制剂,以rhIL-2 200u/ml、PHA 10μg/ml及10％LCM培养,可获得较多的克隆和细胞数。
The clonal culture condition of human NK cells was explored. Human NK cells were expanded from peripheral blood mononuclear cells by cultured with 1000u/ml IL-2 and then purified by complement dependent cytotoxicity(CDC) method to removing T cells. By limited dilution, single NK cell was cultured with allo-PBMC or auto-PBMC( feeder cells) in medium containing rhIL-2, PHA and lymphocyte conditioned medium (LCM). Then cloned cells were collected and identified. Our results indicated that:4 - 16 CD3-CD56 + NK clones(containing 2.35 × 104 cells at most)can be obtained in per 96-well plate. The best cultural condition is :Mitomycin C(25μg/ml) as a feeder cell inhibitor,0.5 or 1 NK cell cultured with feeder cells in medium containing rhIL-2 200u/ml,PHA 10μg/ml and 10%LCM.
出处
《细胞生物学杂志》
CSCD
北大核心
2001年第2期107-110,共4页
Chinese Journal of Cell Biology
基金
国家自然科学基金(No.39870729)
