a b s t r a c tLab-on-a-chip (LOC) devices are broadly used for research in the life sciences and diagnostics and represent a very fast moving field. LOC devices are designed, prototyped and assembled using numerous strategies and materials but some fundamental trends are that these devices typically need to be (1) sealed, (2) supplied with liquids, reagents and samples, and (3) often interconnected with electrical or microelectronic components. In general, closing and connecting to the outside world these miniature labs remain a challenge irrespectively of the type of application pursued. Here, we review methods for sealing and connecting LOC devices using standard approaches as well as recent state-of-the-art methods. This review provides easy-to-understand examples and targets the microtechnology/engineering community as well as researchers in the life sciences.

Performing localized chemical events on surfaces is critical for numerous applications. We earlier invented the microfluidic probe (MFP), which circumvented the need to process samples in closed microchannels by hydrodynamically confining liquids that performed chemistries on surfaces (Juncker et al. Nat. Mater. 2005, 4, 622-628). Here we present a new and versatile probe, the vertical MFP (vMFP), which operates in the scanning mode while overcoming earlier challenges that limited the practical implementation of the MFP technology. The key component of the vMFP is the head, a microfluidic device (∼1 cm(2) in area) consisting of glass and Si and having microfluidic features fabricated in-plane in the Si layer. The base configuration of the head has two micrometer-size channels that inject/aspirate liquids and terminate at the apex which is ∼1 mm(2). In scanning mode, the head is oriented vertically with the apex parallel to the surface with typical spacing of 1-30 μm. Such length scales and using flow rates from nanoliters/second to microliters/second allow chemical events to be performed on surfaces with tens of picoliter quantities of reagents. Before scanning, the head is clipped on a holder for leak-free, low dead volume interface assembly, providing a simple world-to-chip interface. Surfaces are scanned by mounting the holder on a computer-controlled stage having ∼0.1 μm resolution in positioning. We present detailed steps to fabricate vMFP heads having channels with dimensions from 1 μm × 1 μm to 50 μm × 50 μm for liquid localization over areas of 10-10,000 μm(2). Additionally, advanced design strategies are described to achieve high yield in fabrication and to support a broad range of applications. These include particulate filters, redundant aperture architectures, inclined flow-paths that service apertures, and multiple channels to enable symmetric flow confinement. We also present a method to characterize flow confinement and estimate the distance between the head and the surface by monitoring the evolution of a solution of fluorescently labeled antibody on an activated glass surface. This flow characterization reveals regimes of operation suitable for different surface topographies. We further integrate the dispensing of immersion liquid to the vMFP head for processing surfaces for extended periods of time (∼60 min). The versatility of the vMFP is exemplified by patterning fluorescently labeled proteins, inactivation of cells using sodium hypochlorite, and staining living NIH fibroblasts with Cellomics. These applications are enabled by the compact design of the head, which provides easy access to the surface, simplifies alignment, and enables processing surfaces having dimensions from the micrometer to the centimeter scale and with large topographical variations. We therefore believe that ease-of-operation, reconfigurability, and conservative use of chemicals by the vMFP will lead to its widespread use by microtechnologists and the chemical and biomedical communities.

Local interactions between (bio)chemicals and biological interfaces play an important role in fields ranging from surface patterning to cell toxicology. These interactions can be studied using microfluidic systems that operate in the "open space", that is, without the need for the sealed channels and chambers commonly used in microfluidics. This emerging class of techniques localizes chemical reactions on biological interfaces or specimens without imposing significant "constraints" on samples, such as encapsulation, pre-processing steps, or the need for scaffolds. They therefore provide new opportunities for handling, analyzing, and interacting with biological samples. The motivation for performing localized chemistry is discussed, as are the requirements imposed on localization techniques. Three classes of microfluidic systems operating in the open space, based on microelectrochemistry, multiphase transport, and hydrodynamic flow confinement of liquids are presented.

A flexible method to extract more high-quality information from tissue sections is critically needed for both drug discovery and clinical pathology. Here, we present micro-immunohistochemistry (μIHC), a method for staining tissue sections at the micrometre scale. Nanolitres of antibody solutions are confined over micrometre-sized areas of tissue sections using a vertical microfluidic probe (vMFP) for their incubation with primary antibodies, the key step in conventional IHC. The vMFP operates several micrometres above the tissue section, can be interactively positioned on it, and even enables the staining of individual cores of tissue microarrays with multiple antigens. μIHC using such a microfluidic probe is preservative of tissue samples and reagents, alleviates antibody cross-reactivity issues, and allows a wide range of staining conditions to be applied on a single tissue section. This method may therefore find broad use in tissue-based diagnostics and in research.

Point-of-care (POC) immunodiagnostic tests play a crucial role in enabling rapid and correct diagnosis of diseases in prehospital care, emergency, and remote settings. In this work, we present a silicon-based, capillary-driven microfluidic chip integrating two microfluidic modules for the implementation of highly miniaturized immunoassays. Specifically, we apply state-of-the-art microfluidic technology to demonstrate a one-step immunoassay for the detection of the cardiac marker troponin I in human serum using sample volumes of ∼1 μL and with a limit of detection (LOD) of ∼4 ng mL–1 within 25 min. The microfluidic modules discussed here broadly map functionalities found in standard lateral flow assays. We implement a self-coalescence module (SCM) for the controlled reconstitution and delivery of inkjet-spotted and dried detection antibodies (dAbs). This allows for homogeneous dissolution of 1.3 ng of fluorescently labeled dAbs in 416 nL of the sample used for the assay. We also show how to immobilize receptors inside closed microfluidic devices in <30 s using bead lane modules inside which microbeads functionalized with capture antibodies (cAbs) are self-assembled. The resulting bead lane module, with a volume of ∼3 × 10–5 mm3, is positioned across the flow path and holds ∼300 5 μm-diameter microbeads. Altogether, these capillary-driven elements allow for the manipulation of samples and reagents with an unprecedented precision and control, paving the way for the next generation of POC immunodiagnostics.