Chapter 1
Hydrodynamic Flow Confinement Using a Microfluidic Probe

Emmanuel Delamarche, Robert D. Lovchik, Julien F. Cors and Govind V. Kaigala

IBM Research – Zurich, Säumerstrasse 4, 8803 Rüschlikon, Switzerland

1.1 Introduction

Photolithography, the art of patterning surfaces using light projected through an optical mask and chemicals sensitive to light, reached an extraordinary level of sophistication for producing microelectronic components reaching sub-20 nm dimensions on a massive manufacturing scale with extremely high yields. While photolithography had started in the 1960s for fabricating integrated circuits, it essentially remained confined to the structuring and modification of inorganic surfaces and materials. Strong progress on sequencing genomes, an increased understanding of the complexity of cells, tumors, tissues and organs, and emerging work on cell–environment interactions called for new techniques that could tailor biological interfaces and analyze challenging biological specimens. It took, for example, until the 1990s before peptides and oligonucleotides were patterned on glass slides using combinatorial masks and photolithography [1, 2], DNA [3] and protein [4] microarrays were demonstrated, self-assembled monolayers were patterned using soft lithography [5], and various “inks” were deposited on surfaces with nanometer precision using scanning probe methods [6]. All together these techniques are impressive because they can deal with many types of inks, are precise, can cover very large areas, and can be fast and inexpensive. There is, however, a general need for controlling the chemical environment during the deposition of species onto surfaces, the analysis of surfaces, and the study of (bio)interfaces. Controlling the chemical environment here means being able to work with various solutions (biological buffers, culture medium, solvents, etc.) without drying artifacts, potentially at a specific temperature, and being able to change this chemical environment in a flexible manner. The control over the chemical environment on a surface can be achieved, for example, by (i) isolating areas of a surface using microfluidic channels and laminar streams of solutions [7, 8], (ii) applying locally chemicals using a probe [9], or (iii) compartmentalizing chemicals near a surface using nonmiscible liquids [10, 11]. The ability to control a chemical environment on a surface is probably most interesting for biological applications for several reasons: First, investigating the structure and function of proteins, cells, and tissues on surfaces is fundamental. Second, biomolecules and cells are fragile and require appropriate liquid environments such as biological buffers and culture media. Third, cells and tissues are complex heterogeneous systems, and there is a benefit in probing individual cells and particular areas of tissue sections to deal with the complexity of such samples and to use adjacent areas for comparison [12].

Microfluidics represent a powerful approach for interacting with surfaces and studying biological interfaces because the flow of liquids at the microscale is usually laminar and therefore predictable and well defined [13, 14], microfluidic devices can localize chemical processes and environments on surfaces with micrometer precision [15], and microfluidics are conservative of reagents and samples. However, most of microfluidics are closed systems and need either to be sealed on surfaces/samples or samples must be introduced inside microfluidic structures [16, 17]. These challenges can be circumvented by having a device operating in an open space and nearby a surface [18]. Several concepts of “open space” microfluidics have been developed (Figure 1.1). In the example of the FluidFM technology [19], microfluidic channels are integrated to a cantilever for atomic force microscopy, and liquids can be injected or aspirated while scanning a surface. Scanning ion conductance microscopy has also been used to release chemicals using electrophoresis, electroosmosis, and dielectrophoresis from nanopipettes to cells with sub-micrometer precision [20]. These two examples harbor high precision both for scanning and delivery of chemicals but are very dependent on the proximity between the delivering probe and the surface. This requirement can be relaxed by using an aqueous two-phase system. In this case, one phase, which contains the chemicals of interest, can be inserted through the second phase for patterning or interacting with a surface [21]. Finally, a liquid can be localized on a surface using lateral hydrodynamic boundaries formed by a second miscible liquid. This concept is termed hydrodynamic flow confinement (HFC) and can be implemented using the microfluidic probe (MFP) technology [22]. All together, the methods illustrated in Figure 1.1 are powerful for interacting with living cells and tissues and for patterning surfaces with high flexibility on length scales ranging from micrometers to centimeters [18].

Figure 1.1 Local interaction between liquids containing chemicals, biomolecules, and/or cells with a variety of surfaces can be done using scanning noncontact probes. Some of these probes deliver liquids or charged molecules in close proximity to the surface, use nonmiscible liquids to apply a liquid of interest locally to the surface, or use HFC of a liquid inside another one using a multi-aperture microfabricated probe.

This chapter describes the MFP technology in general terms and explains how to design and fabricate MFP heads for implementing HFC of a liquid on a surface. Theoretical descriptions of HFC, applications of the MFP technology, and advanced implementations of HFC as well as alternative approaches for local processing of surfaces are also presented in the following chapters.

1.2 HFC Principle

HFC of a liquid on a surface can be achieved by bringing an MFP head having a face comprising two coplanar microapertures at a distance d above a substrate (Figure 1.2a,b). This distance typically ranges from a few micrometers up to 100 µm, and the resulting gap between the MFP head and the surface is filled with a liquid [22]. This liquid is called the immersion liquid and is typically water, a biological buffer, or a culture medium, depending on the substrate and if proteins or cells are present on the substrate. By injecting and re-aspirating a processing liquid in the gap filled with the immersion liquid through the apertures, the processing liquid flows from one aperture to the other in a laminar regime. This flow is guided by the solid boundaries provided by the substrate and the apex of the MFP head and is directed toward the aspiration aperture together with some of the immersion liquid (Figure 1.2c,d). The immersion liquid is critical for laterally confining the processing liquid in the region beneath the apertures. In addition, HFC can be lost if the ratio of aspiration/injection flow rates (Qa/Qi) reaches a low value. This value essentially depends on d and the distance separating the apertures. As a rule of thumb, Qa should be at least three times Qi [18, 19]. More details on modeling HFC [23, 24] and implementing HFC using more than two apertures [25, 26] are provided in Chapters 2–5.

Figure 1.2 Principle of HFC of a liquid on a substrate using an MFP. (a) A microfabricated head having at least two microapertures is brought close to a surface. (b) When the apex of the MFP head is immersed in an immersion liquid covering a substrate of interest, a processing liquid can be injected and confined on the surface by re-aspirating it together with some of the immersion liquid. (c) The dimensions and spacing of the apertures influence the footprint of the injection liquid on the surface as depicted on this top view. (d) Working on a transparent substrate/sample and having a fluorescent dye in the injection liquid permits direct visualization of the confined liquid using an inverted fluorescence microscope.

The excursion of the processing liquid in space and the consecutive footprint of the processing liquid on a substrate depend on fixed and variable parameters. Fixed parameters are the geometry, lateral dimensions, and spacing of the apertures. Variable parameters are Qi, Qa, d, and the displacement velocities of the MFP head over the substrate (see the following text).

1.3 MFP Heads

The first implementation of HFC using an MFP relied on hybrid heads made from a patterned Si chip bonded to a polydimethylsiloxane (PDMS) layer (Figure 1.3a,b) [22]. Square apertures with lateral dimensions as small as 10 µm were fabricated at the surface of a double-side polished Si wafer using deep reactive ion etching (DRIE). The other side of the wafer was etched to form microchannels and circular cavities. Through-wafer etching then created vias connecting the microchannels to the apertures. A PDMS block comprising through holes was bonded to the Si chip after O2 plasma activation, and glass capillaries were inserted into the holes in the PDMS to provide fluidic connection between computer-controlled syringe pumps and the circular cavities on the Si chip. Structural stability and adhesion between the PDMS and the Si chip were sometimes a problem, limiting the yield of fabrication of such MFP heads. This was solved by making multilayer MFP heads [27]: the lower side of a double-side polished Si wafer comprised microapertures, and the upper side of the wafer was structured with macroscopic channels. A second...