Liquid Crystal Droplet-Based Amplification of Microvesicles that are Shed by Mammalian Cells

摘要

Membrane-derived microvesicles (MVs) shed by cells are being investigated for their role in intercellular communication and as potential biomarkers of disease, but facile and sensitive methods for their analysis do not exist. Here we demonstrate new principles for analysis of MVs that use micrometer-sized droplets of liquid crystals (LCs) to amplify MVs that are selectively captured via antibody-mediated interactions. The influence of the MVs on the micrometer-sized LC droplets is shown to be readily quantified via use of flow cytometry. The methodology was developed using MVs shed by epidermoid carcinoma A431 cells that contain epidermal growth factor receptor (EGFR) as an important and representative example of MVs containing signaling proteins that play a central role in cancer. The LC droplets were found to be sensitive to 10⁶ MVs containing EGFR (relative to controls using isotype control antibody) and to possess a dynamic range of response across several orders of magnitude. Because the 100 nm-sized MVs captured via EGFR generate an optical response in the micrometer-sized LC droplets that can be readily detected by flow cytometry in light scattering mode, the approach possesses significant advantages over direct detection of MVs by flow cytometry. The LC droplets are also substantially more sensitive than techniques such as immunoblotting because the lipid-component of the MVs serves to amplify the antibody-mediated capture of the target proteins in the MVs. Other merits of the approach are defined and discussed in the paper.