Isolation of Circulating Tumor Cells with Acoustophoresis : Towards a biomarker assay for prostate cancer

Abstract: Microfluidics has emerged as an essential approach in the development of novel technological platforms to detect and isolate rare circulating tumor cells (CTCs) in the blood of cancer patients. Micro-scaled fluidic systems offer means to precisely control fluid flow. This enables cell separation when combined with techniques for manipulating cells across fluid streams. Various microfluidic methods have been developed, either using passively generated forces or an applied force field methodology to move cells across streamlines. Acoustophoresis uses ultrasonic standing waves to separate cells and particles in microfluidic channels. An acoustic standing wave field generates acoustic forces that acts on cells and particles based on their individual acoustic properties and forms the basis for the cell separation technology explored in this thesis. In this dissertation, a novel approach for live CTC isolation has been developed. Micron-sized elastomeric particles with negative acoustic contrast were used for negative selection acoustophoresis. The surface of the elastomeric particles was functionalized to bind WBCs through the CD45 antigen, which enabled their transportation to pressure antinodes and facilitated an enrichment of cancer cells at the pressure node. Live cell negative selection acoustophoresis was demonstrated as a proof-of-concept study in paper 1 and was further extended to carry out processing of whole blood by a two-step acoustophoresis method in paper 2. The sample throughput is an important parameter for microfluidic processing of clinical samples, especially for rare cell applications where a larger volume might be required for the detection of target cells. In paper 3, a fluid inertia phenomenon that may compromise cell separation performance at higher flow velocities was discovered. The inertial effects in an acoustofluidic device at increased sample throughputs and its consequences on particle separations were therefore investigated. Through numerical modelling and experimental validation, the main reason for the impaired acoustophoresis separation, at elevated flow rates, was attributed to the formation of a curved fluid boundary between the sample and sheath flow, both at the inlet of the separation channel as well as at the outlet flow splitter.Finally, paper 4 outlines the benchmarking of CTC-acoustophoresis to the FDA cleared CELLSEARCH system in a comparative clinical study. Higher numbers of CTCs were detected after acoustophoretic processing of the patient samples as compared to the CELLSEARCH system. Further studies are currently being conducted to establish the full performance characteristics of CTC-acoustophoresis in the laboratory setting.To conclude, the presented dissertation extends the use of acoustophoresis towards the clinical application of CTC enrichment of live and fixed cells. The aim of establishing an efficient technology that can target the full heterogeneity of the rare tumor cells is essential for the development of novel CTC biomarker assays for metastatic cancer. This dissertation builds towards the goal of an unbiased CTC isolation approach.