Solid Tumor Therapeutics Program, Moores UCSD Cancer Center, UC San Diego, La Jolla, CA 92093, USA.
Methods. 2013 Dec 1;64(2):102-7. doi: 10.1016/j.ymeth.2013.08.008. Epub 2013 Aug 22.
With the emerging interest in personalized medicine, there is strong demand for new technologies for clinical sample interrogation. Exfoliated tumor cells in variety of pathological samples (e.g., blood, bone marrow, urine) could provide invaluable information for diagnosis and prognosis of cancers. Here we describe a detailed method for capture and isolation of tumor cells in medium, blood, or large issue buffy coat using EpCAM-targeted buoyant microbubbles (MBs). Perflorohexane gas lipid shell MBs were prepared with emulsification method and conjugated with antibody as described by us before [25]. The binding of EpCAM-targeted MBs to A549 (human lung carcinoma) and 4T1 (mouse breast carcinoma) cells spiked into BSA/PBS or blood was more than 90%, which was comparable with commercial anti-EpCAM immunomagnetic beads (DynaBeads). Anti-EpCAM MBs efficiently (75-82%) isolated BxPC3 pancreatic tumor cells spiked into medium, blood or a buffy coat, within 15-30 min of incubation. We discuss MB parameters and experimental conditions critical to achieve efficient cells binding and isolation. In conclusion, MB-assisted cell isolation is a promising method for rapid enrichment of cells and biomarkers from biological samples.
随着个性化医疗的兴起，人们对用于临床样本检测的新技术有强烈的需求。在各种病理样本（如血液、骨髓、尿液）中脱落的肿瘤细胞可以为癌症的诊断和预后提供非常有价值的信息。在这里，我们描述了一种使用 EpCAM 靶向浮力微泡（MB）从培养基、血液或大块组织缓冲液中捕获和分离肿瘤细胞的详细方法。全氟己烷气体脂质壳 MB 通过乳化法制备，并按照我们之前的描述与抗体偶联[25]。EpCAM 靶向 MB 与 A549（人肺癌）和 4T1（鼠乳腺癌）细胞的结合率超过 90%，与商业抗 EpCAM 免疫磁珠（DynaBeads）相当。抗 EpCAM MB 可在 15-30 分钟的孵育时间内，有效（75-82%）分离出培养基、血液或缓冲液中掺入的 BxPC3 胰腺肿瘤细胞。我们讨论了对实现有效细胞结合和分离至关重要的 MB 参数和实验条件。总之，MB 辅助的细胞分离是一种从生物样本中快速富集细胞和生物标志物的有前途的方法。
