对比分析了两种卵巢颗粒细胞（granulosa cells, GCs）分离提纯技术，旨在为卵巢储备功能减退（diminished ovarian reserve, DOR）患者提供高效、简便的GCs获取方案。研究收集了160例行体外受精或卵胞质单精子显微注射的DOR患者卵泡液，随机分为改良传统密度梯度离心法组（P/DG组）和颗粒细胞聚集体洗涤法组（W/Agg组），通过纯度鉴定、增殖评估及病理学、形态学与统计学分析。结果表明，两组提取的GCs多聚集成簇，细胞间通过丝状突起相互作用，显微镜下可见脂滴，细胞于24 ~ 48 h开始贴壁生长。流式细胞术显示，W/Agg组悬浮液纯度略低，但总耗时减少，细胞数和活性增加（P < 0.05）。HE染色和FSHR免疫荧光鉴定显示，两组GCs形态完整，呈梭形或星形，W/Agg组GCs生长密度更聚集（P < 0.05）。CCK-8动态监测显示，W/Agg组在第6天增殖高峰更高（P < 0.05）。可见，W/Agg技术在操作效率、细胞数量和活性方面表现更优，尽管初始纯度稍低，但在后续培养中可达到更高的细胞密度和纯度，为优化DOR患者的GCs分离提纯和体外培养技术提供了宝贵的科学依据。

Two techniques for the isolation and purification of ovarian GCs (granulosa cells) were compared and analysed, with the aim of providing an efficient and simple solution for obtaining GCs from patients with DOR(diminished ovarian reserve). 160 follicular fluids from DOR patients undergoing in vitro fertilization or intracytoplasmic sperm injection were collected and randomly divided into P/DG group(modified traditional density gradient centrifugation group) and W/Agg group(granulosa cell aggregation washing group), and the extracted GCs were identified by purity, evaluated for proliferation, and analyzed by pathology, morphology and statistics. The results indicate that GCs extracted from both groups predominantly aggregate into clusters, with cells interacting via filamentous protrusions. Lipid droplets are visible under microscopy, and adherent cell growth begins within 24 ~ 48 h. Flow cytometry analysis reveals that the W/Agg group exhibits slightly lower suspension purity but significantly reduces total processing time while increasing cell yield and viability (P < 0.05). HE staining and FSHR immunofluorescence confirm that GCs in both groups maintain intact morphology, displaying a pike- or star-shaped appearance, with the W/Agg group showing higher cell density and aggregation (P <0.05). CCK-8 dynamic monitoring demonstrates enhanced proliferation in the W/Agg group by day 6 (P < 0.05), with a higher peak activity (P < 0.05). The W/Agg technique proves superior in operational efficiency, cell yield, and viability. Although initial purity is marginally lower, subsequent cultures achieve higher cell density and purity. These findings provide a robust scientific foundation for optimizing GC isolation, purification, and in vitro culture protocols, particularly for patients with DOR.