循环肿瘤细胞 (CTC) 有潜力作为实体瘤的诊断、预后和预测生物标志物。尽管美国食品和药物管理局 (FDA) 批准了 CTC 装置用于治疗各种癌症，但肺癌中 CTC 的稀有性和异质性使得分离和分析它们在技术上具有挑战性，从而阻碍了它们的临床整合。有必要通过对不同 CTC 系统的比较分析来建立共识。本研究旨在使用标准化的掺入方案评估五种技术中的七种不同的 CTC 富集方法：CellMag™（EpCAM 依赖性富集）、EasySep™ 和 RosetteSep™（血细胞去除）以及 Parsortix® PR1 和新的设计 Parsortix® 原型 (PP)（基于尺寸和变形能力的丰富）。还评估了 Parsortix® 系统的细胞收获与盒内染色之间回收率的差异。健康供体血液（5 mL）中加入 100 个荧光标记的 EpCAMhigh H1975 细胞，通过每个系统进行处理，并计算分离效率。 CellMag™ 的回收率最高 (70±14%)，其次是 Parsortix® PR1 盒内染色，而 EasySep™ 的回收率最低 (18±8%)。使用 CellMag™ 和 Parsortix® PR1 盒内染色对 EpCAMmoderate A549 和 EpCAMlow H1299 细胞进行额外的掺入实验。 CellMag™ 对 A549 细胞的回收率显着降低至 35±14%，对 H1299 细胞的回收率显着降低至 1±±1%。然而，Parsortix® PR1 盒内染色显示所有肺癌细胞系的恢复率与细胞表型无关且一致：H1975 (49±±2%)、A549 (47±±10%) 和 H1299 (52±±10%) 。此外，我们证明 Parsortix® PR1 盒内染色方法能够从患者样本中分离异质单一 CTC 和细胞簇。 Parsortix® PR1 盒内染色能够以一致的回收率将不同表型的 CTC 分离为单细胞或细胞簇，被认为是肺癌 CTC 富集的最佳选择，尽管需要进一步优化和验证。© 2024 作者（ s）。约翰·威利出版的《分子肿瘤学》

Circulating tumor cells (CTCs) have potential as diagnostic, prognostic, and predictive biomarkers in solid tumors. Despite Food and Drug Administration (FDA) approval of CTC devices in various cancers, the rarity and heterogeneity of CTCs in lung cancer make them technically challenging to isolate and analyze, hindering their clinical integration. Establishing a consensus through comparative analysis of different CTC systems is warranted. This study aimed to evaluate seven different CTC enrichment methods across five technologies using a standardized spike-in protocol: the CellMag™ (EpCAM-dependent enrichment), EasySep™ and RosetteSep™ (blood cell depletion), and the Parsortix® PR1 and the new design Parsortix® Prototype (PP) (size- and deformability-based enrichment). The Parsortix® systems were also evaluated for any differences in recovery rates between cell harvest versus in-cassette staining. Healthy donor blood (5 mL) was spiked with 100 fluorescently labeled EpCAMhigh H1975 cells, processed through each system, and the isolation efficiency was calculated. The CellMag™ had the highest recovery rate (70 ± 14%), followed by Parsortix® PR1 in-cassette staining, while the EasySep™ had the lowest recovery (18 ± 8%). Additional spike-in experiments were performed with EpCAMmoderate A549 and EpCAMlow H1299 cells using the CellMag™ and Parsortix® PR1 in-cassette staining. The recovery rate of CellMag™ significantly reduced to 35 ± 14% with A549 cells and 1 ± 1% with H1299 cells. However, the Parsortix® PR1 in-cassette staining showed cell phenotype-independent and consistent recovery rates among all lung cancer cell lines: H1975 (49 ± 2%), A549 (47 ± 10%), and H1299 (52 ± 10%). Furthermore, we demonstrated that the Parsortix® PR1 in-cassette staining method is capable of isolating heterogeneous single CTCs and cell clusters from patient samples. The Parsortix® PR1 in-cassette staining, capable of isolating different phenotypes of CTCs as either single cells or cell clusters with consistent recovery rates, is considered optimal for CTC enrichment for lung cancer, albeit needing further optimization and validation.© 2024 The Author(s). Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.