Pipetting-driven microfluidic immunohistochemistry to facilitate enhanced immunoreaction and effective use of antibodies

Abstract

Immunohistochemistry (IHC), which has been used to detect antigens in cells of a tissue section using an immunoreaction between an antibody and an antigen, is a practical tool for identifying the type and stage of diseases in cancer diagnosis and scientific research. However, conventional IHC requires long, laborious process times and high costs. Although microfluidic IHC platforms have been developed to overcome these limitations, the application of microfluidic IHC in real-world environments is still limited due to the additional equipment needed to operate the microfluidic systems. In addition, continuous flow in a microfluidic channel leads to a waste of unbound antibodies. In this study, we demonstrate a novel and easy-to-use microfluidic IHC platform operated only using a manual pipette that is commonly available in research laboratories or hospitals. No other device such as a pump or a controller is required to operate our system. Bidirectional flows of the antibody solution in a microfluidic device are induced by repetitive manual pipetting which facilitates the enhanced antigen-antibody reaction and enables the effective use of a limited amount of antibody. When breast cancer cell and tissue sections are reacted with antibodies using our platform, pipetting for less than 2 min is sufficient to obtain immunostaining results without damaging the sample. The staining intensity by our method is similar to that of the sample stained for 1 h by a conventional batch process. We believe that this pipetting-based approach to the operation of a microfluidic system allows end users to use microfluidic IHC more conveniently and easily in real-world environments.

Rapid and efficient western blot assay by rotational cyclic draining and replenishing procedure

“…This phenomenon can be implemented by various strategies, for example, simply by flipping the probe incubation chamber continuously or by tilting the container of probe solution back and forth . It can also be achieved by lateral flow of probe solution in microfluidic device generated by the negative pressure of pipetting . Multiple studies demonstrated that CDR shortened assay time from hours to minutes in common surface bioassays like ELISA and immunofluorescence microscopy .…”

Acoustofluidic devices controlled by cell phones

“…As explained previously, micropumping and micromixing both enable precise, controlled, and rapid reactions, assays, and diagnostics tests. 8,37,63,64 While each of these functions is critical for many microfluidic applications, they remain discrete innovations. Having developed the sharp-edge based technology necessary to achieve both functions separately, herein we also demonstrated that by combining the design of sharp-edge structures for micropumping and micromixing together, concurrent pumping and mixing can be achieved on a single acoustofluidic chip.…”

Microfluidic on-chip immunohistochemistry directly from a paraffin-embedded section

“…For the IHC assay, we used a pressure-based reversible sealing method to seal the microfluidic channel by applying pressure on the chip that is in contact with the section slide. 19,24 However, the UV-curable PUA used in this work also has a rigid property after curing and causes a sealing problem. Since a typical microfluidic chip has a flat surface, it has a large contact area with the sample, thereby lowering the pressure applied to the channel surface.…”

“…13,14 These advantages have led to the development of various microfluidic IHC platforms that are capable of performing complex assays, such as multiplexed biomarker screening on breast cancer tissue, 15,16 precise spatial control of immunostaining, 17,18 and enhanced immunoreaction in a reduced process time. 19 An accurate IHC analysis for biomarker quantification was also demonstrated using a quantum-dot based microfluidic double-staining method, 20,21 a microfluidic tissue processor for formalin-fixed sections, 22 and frozen sections, 23 and standardization of immunostaining quality. 24 Although these microfluidic techniques have contributed to expand the potential of the IHC process, there are still a limited number of applications.…”