Circulating tumor cells (CTCs) are the cells that are shed from a primary tumor into the vasculature and circulate in the bloodstream. CTCs may trigger cancer metastasis, which leads to most cancer-related deaths. CTCs are widely studied due to their value in cancer diagnosis, prognosis, and oncology studies. The major challenges with CTCs lie in their extremely low concentration in blood, thus requiring an effective enriching system to enable downstream analyses. The immunomagnetic assay has proved to be a promising CTC detection tool with high sensitivity and throughput. Key factors related to the immunomagnetic assay include the capture rate, which indicates the sensitivity, and distributions of target cells after capture, which impact the cell integrity and other biological properties. In this dissertation, we build a sedimentation model, a partial viscosity model, and a cell-tracking model to address the principle of the immunomagnetic cell separation. We examine the channel orientations and determine the favorable inverted condition. In addition, we develop a micromagnet approach to modulate the in-channel magnetic field toward enhanced cell detection and distribution. Through numerical studies, we calculate the magnetic field generated by the thin-film micromagnets, determine its effective ranges, and demonstrate its value in optimizing cell distribution. In the experimental demonstration, we present two types of micromagnets based on e-beam Ni deposition and inkjet printing technology, respectively. In the screening experiments, the Ni micromagnet integrated system achieves over 97% capture rate. It shows a 14% increase in capture rate, and a 14% improvement in distribution uniformity compared with plain slides. We also successfully isolate CTCs from metastatic cancer patients with the micromagnet assay. The inkjet-printed patterns yield a similarly high capture rate of 103%. With the pixel permanent magnet array, the inkjet patterns further increase the distribution uniformity for 20%. The proposed models lay the theoretical foundations for future modification of the immunomagnetic assay, and the micromagnet-integrated system provides a promising tool for translational applications in cancer diagnose and clinical cancer management. / text

Circulating tumor cells (CTCs) are known to escape from the primary tumor site and may settle down at the distant organ to grow a second tumor. CTCs are one of causes initiating carcinoma metastasis. Detection of CTCs has been considered to be valuable for cancer management, including diagnosis, prognosis, and clinical treatment management. However, efficient isolation, enumeration, characterization, and genetic analysis of CTCs in whole-blood samples from cancer patients are very challenging due to their extremely low concentration and rare nature (per CTC in blood cells is 1:106–109). With the increasing worldwide death rate associated with cancer, there is a desperate demand for a high-sensitivity, high-throughput, and low-cost detection and separation system. My doctoral research focused on the design and fabrications of the screening system for the detection of CTCs with further analysis of captured CTCs, such as immunofluoresce staining and fluorescence in-situ hybridization (FISH). The distinct significance of this research is that the development of the computer-controlled rotational holder with a series of six inverted microfluidic chips reduced the cost by significantly reducing the consumption of magnetic carriers (25% of the consumed amount used in the commercial CellSearch® system), increasing the capture efficiency by manipulating the blood sedimentation in the microchannel, enhancing the system stability by integrating the micromagnets on the plain glass slide substrate, and achieving high throughput because of the high flow rate (2.5 mL/hr) and large screening volume (screening up to six chips in parallel with each containing 2.5 mL of blood). Immunofluorescence staining and the FISH method have been performed to prove the capability of the system. In addition, the system has been successfully applied for patient samples screening. The incorporation of micromagnets has demonstrated that micromagnets provide localized magnetic forces to scatter the target cancer cells and free nanoparticles throughout the whole channel substrate to increase the channel space usage by 13%. Four cancer cell lines, including COLO 205 (colorectal cancer), SK-BR-3 (breast cancer), MCF-7 (breast cancer), and PC3 (prostate cancer), were spiked in blood samples from healthy donors to verify high capture efficiency of the developed system. On average, over a 97% capture rate was demonstrated for all cell lines. Moreover, the developed screening system has been successfully screened over 40 patient samples, including metastatic lung cancer, breast cancer, prostate cancer, and colorectal cancer. After capture of CTCs, immunofluorescence staining was used to identified the captured cancer cells and the FISH method was performed to characterize the isolated cancer cells by studying the gene expression of CTCs from breast cancer. The proposed automated immunomagnetic microchip-based screening system shows high capture efficiency (average 97% for three spiked cell lines), high throughput (15 mL of blood sample per screening), high sensitivity, high specificity, and low nanoparticle consumption (75% less than CellSearch® system). The screening system provides great promise as a clinical tool for early cancer diagnosis, diagnosis, personalized therapy, and treatment monitoring. / text

L'auteur n'a pas fourni de résumé en français / Nowadays in medicine and biotechnology a wide range of applications involves magnetic micro/nano-object manipulation including remote control of magnetic beads, trapping of drug vectors, magnetic separation of labelled cells and so on. Handling and positioning magnetic particles and elements functionalized with these particles has greatly benefited from advances in microfabrication. Indeed reduction in size of the magnet while maintaining its field strength increases the field gradient. In this context, arrays made of permanent micromagnets are good candidates for magnetic handling devices. They are autonomous, suitable for integration into complex systems and their magnetic action is restricted to the region of interest.In this thesis we have elaborated an original approach based on AFM and MFM for quantitative study of the magnetic force and associated force gradients induced by TMP micromagnet array on an individual magnetic micro/nano-object. For this purpose, we have fabricated smart MFM probes where a single magnetic (sub)micronic sphere was fixed at the tip apex of a non-magnetic probe thanks to a dual beam FIB/SEM machine equipped with a micromanipulator.Scanning Force Microscopy conducted with such probes, the so-called Magnetic Particle Scanning Force Microscopy (MPSFM) was employed for 3D mapping of TMP micromagnets. This procedure involves two main aspects: (i) the quantification of magnetic interaction between micromagnet array and attached microsphere according to the distance between them and (ii) the complementary information about micromagnet array structure. The main advantage of MPSFM is the use of a probe with known magnetization and magnetic volume that in combination with modelling allows interpreting the results ably.We conducted MPSFM on TMP sample with two types of microparticle probes: with superparamagnetic and NdFeB microspheres. The measurements carried out with superparamagnetic microsphere probes reveal attractive forces (up to few tens of nN) while MFM maps obtained with NdFeB microsphere probes reveal attractive and repulsive forces (up to one hundred of nN) for which the nature of interaction is defined by superposition of microsphere and micromagnet array magnetizations. The derived force and its gradient from MFM measurements are in agreement with experiments on microparticle trapping confirming that the strongest magnetic interaction is observed above the TMP sample interfaces, between the areas with opposite magnetization. Thanks to 3D MFM maps, we demonstrated that intensity of magnetic signal decays fast with the distance and depends on micromagnet array and microsphere properties.Besides the magnetic interaction quantification, we obtained new information relevant to TMP sample structure: we observed and quantified the local magnetic roughness and associated fluctuations, in particular in zones of reversed magnetization. The variation of detected signal can reach the same order of magnitude as the signal above the micromagnet interfaces. These results complete the experiments on particle trapping explaining why magnetic microparticles are captured not only above the interfaces, but also inside the zones of reversed magnetization.Quantitative measurements of the force acting on a single (sub)microsphere associated to the modelling approach improve the understanding of processes involved in handling of magnetic objects in microfluidic devices. This could be employed to optimize the parameters of sorting devices and to define the quantity of magnetic nanoparticles required for labelling of biological cells according to their size. More generally these experimental and modelling approaches of magnetic interaction can meet a high interest in all sorts of applications where a well-known and controlled non-contact interaction is required at micro and nano-scale.