Abstract
Technologies for the detection and isolation of circulating tumor cells (CTCs) are essential in liquid biopsy, a minimally invasive technique for early diagnosis and medical intervention in cancer patients. A promising method for CTC capture, using an affinity-based approach, is the use of functionalized hydrogel microparticles (MP), which have the advantages of water-like reactivity, biologically compatible materials, and synergy with various analysis platforms. In this paper, we demonstrate the feasibility of CTC capture by hydrogel particles synthesized using a novel method called degassed mold lithography (DML). This technique increases the porosity and functionality of the MPs for effective conjugation with antibodies. Qualitative fluorescence analysis demonstrates that DML produces superior uniformity, integrity, and functionality of the MPs, as compared to conventional stop flow lithography (SFL). Analysis of the fluorescence intensity from porosity-controlled MPs by each reaction step of antibody conjugation elucidates that more antibodies are loaded when the particles are more porous. The feasibility of selective cell capture is demonstrated using breast cancer cell lines. In conclusion, using DML for the synthesis of porous MPs offers a powerful method for improving the cell affinity of the antibody-conjugated MPs.
Highlights
During the cancer metastatic process, malignant cells detach from the primary tumor, enter the blood stream, and relocate at a distal site
We demonstrated the feasibility of selectively capturing cancer cells by hydrogel particles synthesized using our degassed mold lithography (DML) technique
The DML technique enables high throughput synthesis of the described hydrogel particles without the limitations of short UV exposure and microfluidic challenges associated with conventional approaches, such as stop flow lithography (SFL)
Summary
During the cancer metastatic process, malignant cells detach from the primary tumor, enter the blood stream, and relocate at a distal site. Of the two general approaches, the most popular approach for CTC capture is affinity-based techniques [8,24,25,26,27,28,29], due to their high performance in CTC detection and isolation They employ a wide range of specific monoclonal antibodies, lectins, and aptamers to monitor the various types of molecules on the cell surface. To increase the porosity of a PEG hydrogel, a low content of cross-linking monomer is used with a high content of porogen in the prepolymer [36,37,38] This synthesis condition is challenging to apply using the previously described SFL fabrication technique [30], as SFL has a short UV exposure time to facilitate its high throughput and microfluidic system stability, which produces particles with poor network integrity and, occasionally, even failure in the formation of particle synthesis. As proof of concept, optimized hydrogel particles were applied in a cell line assay to demonstrate that target cell capture can be achieved through selective antibody functionalization
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