Circulating tumor cells (CTCs) are cancer cells shredded from either a primary tumor or a metastatic site and circulate in the blood as the potential cellular origin of metastasis. By detecting and analyzing CTCs, we will be able to noninvasively monitor disease progression in individual cancer patients and obtain insightful information for assessing disease status, thus realizing the concept of “tumor liquid biopsy”. However, it is technically challenging to identify CTCs in patient blood samples because of the extremely low abundance of CTCs among a large number of hematologic cells. In order to address this challenge, our research team at UCLA pioneered a unique concept of “NanoVelcro” cell-affinity substrates, in which CTC capture agent-coated nanostructured substrates were utilized to immobilize CTCs with remarkable efficiency. Four generations of NanoVelcro CTC assays have been developed over the past decade for a variety of clinical utilities. The 1st-gen NanoVelcro Chips, composed of a silicon nanowire substrate (SiNS) and an overlaid microfluidic chaotic mixer, were created for CTC enumeration. The 2nd-gen NanoVelcro Chips (i.e., NanoVelcro-LMD), based on polymer nanosubstrates, were developed for single-CTC isolation in conjunction with the use of the laser microdissection (LMD) technique. By grafting thermoresponsive polymer brushes onto SiNS, the 3rd-gen Thermoresponsive NanoVelcro Chips have demonstrated the capture and release of CTCs at 37 and 4 °C respectively, thereby allowing for rapid CTC purification while maintaining cell viability and molecular integrity. Fabricated with boronic acid-grafted conducting polymer-based nanomaterial on chip surface, the 4th-gen NanoVelcro Chips (Sweet chip) were able to purify CTCs with well-preserved RNA transcripts, which could be used for downstream analysis of several cancer specific RNA biomarkers. In this review article, we will summarize the development of the four generations of NanoVelcro CTC assays, and the clinical applications of each generation of devices.

循环肿瘤细胞（CTC）是从原发肿瘤或转移部位切碎的癌细胞，并在血液中循环，作为潜在的转移细胞起源。通过检测和分析CTC，我们将能够无创地监测单个癌症患者的疾病进展，并获得有洞察力的信息以评估疾病状态，从而实现“肿瘤液体活检”的概念。但是，由于大量血液细胞中CTC的丰度极低，因此在患者血液样本中鉴定CTC在技术上具有挑战性。为了应对这一挑战，我们在UCLA的研究团队率先提出了“ NanoVelcro”细胞亲和性底物的独特概念，其中利用CTC捕获剂涂层的纳米结构化底物以惊人的效率固定了CTC。在过去的十年中，已经开发出了四代的NanoVelcro CTC测定法，用于各种临床用途。创建了由硅纳米线基板（SiNS）和覆盖的微流体混沌混合器组成的第一代NanoVelcro芯片，用于CTC枚举。基于聚合物纳米基质的第二代NanoVelcro芯片（即NanoVelcro-LMD）是结合激光显微切割（LMD）技术开发的，用于单CTC分离。通过将热敏性聚合物刷接枝到SiNS上，第三代热敏性NanoVelcro芯片分别展示了在37°C和4°C下捕获和释放CTC的能力，从而可以快速纯化CTC，同时保持细胞活力和分子完整性。用硼酸接枝的导电聚合物基纳米材料在芯片表面上制备，第四代NanoVelcro芯片（Sweet芯片）能够用保存完好的RNA转录本纯化四氯化碳，可用于下游分析多种癌症特异性RNA生物标记物。在这篇综述文章中，我们将总结四代NanoVelcro CTC分析的发展以及每一代设备的临床应用。