The microfluidic probe (MFP) is a non-contact technology that applies the concept of hydrodynamic flow confinement within a small gap to eliminate the need for closed conduits, and thus overcomes the conventional microfluidic “closed system” limitations. It is an open-space microfluidic concept, where the fluidic delivery mechanism is physically decoupled from the target surface to be processed such as tissue slices or cell culture in Petri dishes. Typically, MFPs are manufactured using complex photolithography-based microfabrication procedures that limits innovation in MFPs’ design and integration. Recently, we showed that 3D printing can be utilized for rapid microfabrication of MFPs, where MFPs can be manufactured with built-in components such as reservoirs, fluidic connectors, and interfaces to the XYZ probe holder. 3D printing brings flexibility in MFP design, where different configurations and aperture arrangements can be considered. Currently, we are developing advanced MFPs that are integrated with other technologies and targeting applications in dielectrophoretic-based cell separation, immuno-based cell capture for isolating circulating tumor cells from blood samples, and efficient and selective single cell electroporation. In this invited paper, we highlight several MFP technologies we are developing.
This work presents the development of a micro-electro-fluidic probe (MeFP) platform as an affordable and flexible microfluidic tool for the transfection of single cells via electroporation. The platform constitutes of a 3D printed MeFP -- gold-coated microfluidic probe (MFP) with an array of pin shaped microelectrodes integrated on its tip -- and an ITO coated cell culture substrate. This setup, and submicron feature size of the MeFP, allows for a selective exposure of the targeted cell to both the electric field and hydrodynamic flow confinement (HFC) of an intercalating agent, to demonstrate transmembrane molecule delivery through electroporation. Results show successful transfer of propidium iodide (PI) through the membranes of single HeLa cells with an applied DC rectangular pulse– a proof-of-concept for MeFP’s application in delivering nucleic acids into eukaryotic cells (transfection). By adjusting the size of the HFC (varying injection and aspiration flow ratio), we show that the cell target area can be dynamically increased from the single cell footprint, to cover multiple cells. Finite Element model show that even with such low applied voltages (0.5- 3Vpk-pk), the electric field generated reach the reversible electroporation threshold. These results demonstrate the MeFP as an advancement to the currently available transfection technologies for gene therapy; delivery of DNA vaccines, in vitro fertilization, cancer treatment, regenerative medicine, and induced pluripotent stem (iPS) cells.
Effective capture of cancer cells from whole peripheral blood samples, i.e. circulating tumor cells (CTCs), is still an existing limitation for liquid biopsy-based diagnostics. The well-established closed-channel herringbone micro-mixers are one of the widely adopted methods for isolating CTCs based on antigen-antibody interaction. However, they are known to be associated with several drawbacks, such as limited capture areas within the channels, restricted access to isolated cells, difficulties to achieve multiplexed antibody capture assays for immuno-phenotyping, and limited postprocessing possibilities. To tackle these issues, we developed a novel microfluidic probe (MFP) that is integrated with herringbone micro-mixers on its tip surface (HMFP). The tip surface was designed with 2-slitted apertures, one for injecting the cell suspension and the other for performing high flow rate aspiration to confine the flow. The herringbone mixing elements were distributed in-between the apertures for micro-mixing that enhances the CTCs capture on the antibodies-coated bottom glass surface. Unlike the closed herringbone chips, the functionalized bottom glass surface was kept large given the capacity for the MFP to work in scanning mode, and so it prevented cell capture saturation effect. Our MFP design and experimental setup showed a cell capture efficiency of 59-81% with flow rates of 0.6-2.4 mL/h. The capture of CTCs in an open microfluidic system allows for easy post-process and CTC analysis, such as single cell drug testing and mechano-phenotyping using atomic force microscopy.